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GlES  FELLOWSHIP 

in  Biolosical  Chemistry 


Digitized  by  the  Internet  Archive 

in  2010  with  funding  from 

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http://www.archive.org/details/laboratorymanual1915aute 


THE  DETECTION  OF 
POISONS  AND  POWERFUL  DRUGS 


AUTENRIETH— WARREN 


LABORATORY  MANUAL 

FOR 

The  Detection  of  Poisons 

AND 

Powerful  Drugs 


BY 

DR.  WILHELM  AUTENRIETH 

PROFESSOR   IN  THE   UNIVERSITY  OF   FREIBURG   i.   B. 


AUTHORIZED  TRANSLATION 

OF    THE 

COMPLETELY  REVISED  FOURTH  GERMAN  EDITION 


BY 

WILLIAM  H.  WARREN,  Ph.D. 

PROFESSOR  OF  CHEMISTRY    IN    WHEATON   COLLEGE 


WITH  25  ILLUSTRATIONS 


PHILADELPHIA 

P.   BLAKISTON'S   SON   &   CO. 

1012  WALNUT  STREET 


Copyright,  1915,  by  P.  Blakiston's  Son  &  Co. 


ni5* 


THE. MAPLE. PRESS. YORK- PA 


AUTHOR'S  PREFACE 


Additional  matter  in  "Detection  of  Poisons"  has  made  the 
fourth  edition  considerably  larger  than  the  third.  The  seven 
chapters  now  comprised  in  the  book  have  been  entirely  revised, 
but  the  first  three  chapters  remain  unchanged  in  arrangement. 
Chapter  I  treats  of  poisons  volatile  with  steam.  Organic 
poisons,  especially  the  alkaloids,  form  the  subject  of  Chapter  II. 
Hydrastine  and  veronal,  introduced  into  this  chapter  for  the 
first  time,  have  been  incorporated  into  the  Stas-Otto  process. 
Chapter  III  deals  with  metallic  poisons. 

The  toxic  substances  included  in  Chapter  IV  find  no  place  in 
the  three  groups  just  mentioned.  As  they  seldom  appear  in 
toxicological  examinations,  they  are  of  theoretical  rather  than 
of  practical  significance.  The  following  members  of  this  group 
have  been  introduced  for  the  first  time,  namely,  cantharidin, 
cytisine,  ergot,  papaverine,  pilocarpine,  saponin  substances, 
solanine,  thebaine,  and  the  toxalbumins,  ricin,  abrin  and  crotin. 

Chapter  V  has  to  do  with  special  qualitative  and  quantitative 
methods  such  as  the  quantitative  estimation  of  phosphorus  in 
phosphorated  oils;  the  electrolytic  detection  and  estimation  of 
arsenic;  the  biological  test  for  arsenic;  the  destruction  of  organic 
matter  and  detection  of  arsenic  by  A.  Gautier  and  G.  Locke- 
mann;  Karl  Th.  Morner's  estimation  of  minute  quantities  of 
arsenic;  methods  of  estimating  alkaloids  by  H.  Matthes,  H. 
Thoms  and  A.  H.  Gordin.  This  chapter  also  includes  A.  J.  J. 
Vandevelde's  estimation  of  the  toxic  action  of  organic  com- 
pounds by  means  of  blood  haemolysis. 

Chapter  VI  takes  up  the  estimation  of  alkaloids  and  other 
active  principles  in  raw  materials  (drugs)  and  in  their  prepara- 
tions. Pharmacopoeial  as  well  as  other  estimations  such  as 
that  of  nicotine  in  tobacco,  caffeine  in  tea,  coft'ee,  kola  prepara- 
tions, etc.,  pilocarpine  in  jaborandum  leaves,  piperine  in  pepper, 

V 


Vi  AUTHOR  S   PREFACE 

solanine  in  potatoes,  and  theobromine  in  cacao  and  its  prepara- 
tions have  been  included.  The  author  has  endeavored  to  treat 
these  subjects  as  thoroughly  as  possible. 

Chapter  VII  describes  the  methods  employed  in  detecting 
carbon  monoxide  in  blood,  in  recognizing  blood  itself  in  stains 
and  in  differentiating  human  from  animal  blood. 

The  new  edition,  though  more  comprehensive  than  the  last 
in  its  scope,  has  lost  nothing  in  clearness  because  of  the  rear- 
rangement of  subject  matter.  Beginners  will  probably  confine 
their  attention  to  the  first  three  chapters.  Students  of  phar- 
macy will  undoubtedly  add  Chapter  VI  which  deals  with  drug 
assaying.  The  other  chapters  are  designed  more  especially  for 
those  who  wish  to  become  better  acquainted  with  toxicological 
procedures. 

Descriptions  of  syntheses  of  organic  drugs  such  as  acetanilide, 
antipyrine,  phenacetine,  pyramidone,  salicylic  acid,  sulphonal 
and  veronal  allow  the  student  to  review  the  methods  employed 
in  connection  with  laboratory  work.  Structural  formulae  of 
alkaloids  and  their  cleavage  products  have  been  given  only  when 
they  have  been  definitely  determined  or  shown  to  be  highly 
probable.  By  introducing  this  specific  information  the  author 
hopes  to  stimulate  the  student's  interest  in  alkaloidal  chemistry 
which  has  become  so  important  within  recent  years. 

More  advanced  students  will  find  in  fine  print  brief  state- 
ments about  the  poisonous  action  of  the  better  known  physiologi- 
cally active  substances  as  well  as  their  distribution  in  and  elimi- 
nation from  the  human  organism.  Repeated  references  to 
larger  treatises  upon  toxicology,  especially  to  R.  Kobert's 
"Intoxikationen",  have  been  made.  Numerous  citations  of 
literature  enable  the  student  to  consult  original  articles  for  fuller 
information. 

The  translation  of  the  third  and  fourth  editions  into  English 
and  Spanish  and  the  proposed  translation  of  the  fourth  edition 
into  Italian  indicate  that  colleagues  in  other  countries  have 
favorably  received  this  work. 

WiLHELM  AUTENRIETH. 

Freiburg  in  Baden. 


TRANSLATOR'S  PREFACE 


The  introduction  of  new  matter  and  certain  rearrangements  of 
the  text  make  the  fourth  edition  of  ** Detection  of  Poisons" 
quite  different  from  the  last.  Without  exception  these  changes 
have  added  to  the  value  of  the  book  not  only  as  a  laboratory 
manual  for  students  but  as  a  guide  for  those  wishing  to  make 
practical  use  of  the  procedures  described.  The  translation  fol- 
lows the  German  as  closely  as  is  consistent  with  clearness.  As 
in  the  translation  of  the  third  edition,  the  methods  from  the 
German  Pharmacopoeia  remain  unchanged. 

Aside  from  the  introduction  of  a  few  substances  that  do  not 
appear  in  the  earlier  editions  and  the  addition  of  new  methods, 
the  general  plan  of  the  first  three  chapters  is  the  same  as  that 
of  the  last  edition.  Believing  that  the  subject  of  so-called 
normal  arsenic  in  Chapter  III  is  not  presented  in  the  German 
edition  at  suihcient  length  to  do  full  justice  to  both  sides  of  the 
question,  the  translator  upon  his  own  responsibility  has  under- 
taken to  give  a  complete  statement  of  the  case  with  citations  of 
the  principal  authorities. 

Most  that  is  new  in  the  book  appears  in  Chapter  V  which 
treats  of  special  methods  of  analysis.  In  addition  to  the  meth- 
ods given  in  the  German  edition,  the  translator  has  thought  it 
worth  while  to  introduce  the  quantitative  estimation  of  arsenic 
and  antimony  by  the  Gutzeit  method  as  worked  out  by  the  late 
Professor  Sanger.  The  procedure  is  so  simple  that  it  may  appeal 
to  some  chemists  as  a  desirable  substitute  for  the  more  com- 
monly used  Marsh-Berzelius  test.  Otherwise  the  translation 
has  not  departed  from  the  German  text  in  any  essential  way. 

William  H.  Warren. 

Norton,  Massachusetts. 


vu 


CONTENTS 

Paob 

Author's  Preface iii 

Translator's  Preface      v 

Introduction I 

CHAPTER  I 

Tests  for  Phosphorus  and  Other  Poisons  Volatile  with  Steam  from 

Acid  Solution 

Phosphorus 5 

Scherer's  test;  Mitscherlich's  test;  Blondlot  and  Dusart's  test; 
(a)  in  the  Fresenius-Neubauer  apparatus,  (b)  in  the  Hilger- 
Nattermann  apparatus;  Detection  of  phosphorous  acid;  Phos- 
phorus in  phosphorated  oils;  Detection  and  quantitative 
estimation  by  the  Mitscherlich-Scherer  method;  Metabolism 
in  phosphorus  poisoning. 

Further  Examination  of  the  Distillate 

Hydrocyanic  Acid 19 

Physiological  action ;  Preliminary  test;  Detection;  Quantitative 
estimation;  Detection  in  presence  of  potassium  ferrocyanide; 
Mercuric  cyanide;  Mercuric  cyanide  in  presence  of  potassium 
ferrocyanide. 

Carbolic  Acid 26 

Action  and  fate  in  the  organism;  Detection;  Quantitative  esti- 
mations; I.  Gravimetrically;  2.  Volumetrically  (Beckurts- 
Koppeschaar) ;  3.  Volumetrically  (J.  Messinger-G.  Vortmann); 
Estimation  in  urine;  Carbolic  acid  in  presence  of  aniline. 

Chloroform 35 

Behavior  in  the  human  organism;  Distribution  in  the  cadaver; 
Detection;  Quantitative  estimation  in  cadaveric  material. 

Chloral  Hydrate 38 

Detection;  Action  and  fate  in  the  human  organism;  Quanti- 
tative estimation  in  blood  and  tissues. 

Iodoform 41 

Detection. 

Nitrobenzene      42 

Toxic  action;  Detection. 

Aniline 44 

Toxic  action;  Detection. 

Carbon  Disulphide 46 

Toxic  action;  Detection;  Quantitative  estimation  of  carbon 
disulphide  vapor  in  air. 

ix 


X  CONTENTS 

Page 
Ethyl  Alcohol 49 

Fate  in  the  human  organism;  Detection. 
Acetone 51 

Occurrence  in  urine;  Detection;  Acetone  in  presence  of  ethyl 

alcohol;  Detection  in  urine. 

Bitter  Almond  Water  and  Benzaldehyde 53 

Synopsis  of  Group  I  (Chapter  I)  55 

CHAPTER  II 

Detection  of  those  Organic  Substances  which  are  not  Volatile  with 
Steam  from  Acid  Solution 

Stas-Otto  process.         59 

A.  Examination  of  Ether  Extract   of  the  Aqueous  Tartaric 

Acid  Solution 59 

PiCROTOXIN 61 

Detection  in  beer. 

Colchicin 64 

Picric  Acid 65 

Action  and  Elimination;  Detection, 
Acetanilide 68 

Action;  Detection;  Examination  of  acetanilide  urine. 
Phenacetine 70 

Preparation;  Detection. 
Salicylic  Acid      72 

Detection;  Quantitative  estimation;  Detection  in  urine. 
Veronal 75 

Preparation;  Physiological  action;  Detection;  In  urine. 
Antipyrine 78 

Preparation;  Detection;  In  urine. 
Caffeine 79 

Fate  in  human  metabolism;  Detection. 

B.  Examination  of  Ether  Extract  of  the  Aqueous  Alkaline 

Solution '. 81 

CONIINE 85 

Nicotine 86 

Physiological  action;  Reactions. 

Aniline 89 

Veratrine 89 

Preparation  of  crystalline  and  water  soluble  veratrine;  Con- 
stitution; Reactions. 

Strychnine 92 

Physiological  action;  Detection;  Detection    of    strychnine    in 
presence  of  brucine. 

Brucine 96 

Atropine 98 

Constitution ;  Reactions. 


CONTENTS  XI 

Pace 
HOMATROPINE lOI 

Cocaine loi 

Constitution;  Behavior  in  the  animal  organism ;  Detection. 

Physostigmine 105 

Codeine 106 

Narcotine 108 

Constitution;  Detection. 
Hydrastine 112 

Preparation;  Constitution;  Reactions. 
Quinine 114 

Constitution;  Detection. 

Caffeine 118 

Antipyrine 118 

Detection  in  urine. 
Pyramidone 119 

Preparation;  Behavior  in  the  organism;  Detection. 

C.  Examination  of  Ether  Extract  and  of  Chloroform  Extract 
OF  THE  Solution  Alkaline  with  Ammonia 

a.  Ether  Extract 122 

Apomorphine 122 

/3.  Chloroform  Extract 124 

Preliminary  test  for  morphine;  Purification  of  crude  morphine. 

Morphine 126 

Constitution;  Detection;  Behavior  in  the  animal  organism. 

Narceine 131 

Constitution;  Reactions. 

Synopsis  of  Group  II  (Chapter  II)       134 

CHAPTER  III 

Examination  for   Metallic  Poisons 

Fresenius-v.  Babo  Method  of  destroying  organic  matter  ....  141 

Destruction  of  organic  matter  with  free  chloric  acid 144 

C.  Mai's  Method  of  destroying  organic  matter 145 

Precipitation  with  hydrogen  sulphide 145 

Metallic  Poisons  I:  Examination  of  that  portion  of  the  h^'drogen 
sulphide  precipitate  soluble  in  ammonia-ammonium  sulphide. 

Arsenic 149 

Marsh-Berzelius  method;  Fresenius-v.  Babo  method;  Betten- 
dorff's  arsenic  test;  Gutzeit's  arsenic  test. 

Antimony,  Tin,  Copper 156 

Metallic  Poisons  II :  Examination  of  that  portion  of  the  hydrogen 

sulphide  precipitate  insoluble  in  ammonium  sulphide     ....  158 

Mercury,  Lead,  Copper,  Bismuth,  Cadmium 158 

Metallic  Poisons  III:  Examination  for  Chromium  and  Zinc    .    .    .  161 

Zinc ' 161 


Xll  CONTENTS 

Page 

Chromium 162 

Metallic  Poisons  IV:  Examination  for  Barium,  Lead  and  Silver  of 
the  insoluble  residue  left  on  treatment  with  potassium  chlorate 

and  hydrochloric  acid 163 

Synopsis  of  Group  III  (Chapter  III) 164 

The  Action  of  Heavy  Metals 165 

Fate,  Distribution  and  Elimination  of  Metals  in  the  body  .  166 

CHAPTER  IV 

1.  Examination  for  those  Poisons  which  do  not  belong  to  the  Three 

Main  Groups  of  Poisons 

Mineral  Acids 

Hydrochloric  Acid 176 

Nitric  Acid 177 

Sulphuric  Acid 179 

Sulphurous  Acid 181 

Oxalic  Acid 182 

Toxic  action;  Distribution  in  the  organism;  Detection. 

Detection  of  Free  Alkalies 

Potassium  Hydroxide,  Sodium  Hydroxide,  Ammonia 185 

Potassium  Chlorate 187 

Toxic    action;  Detection;  Quantitative    estimation;  Behavior 

during  putrefaction;  Detection  in  meat. 

Examination  for  Santonin,  Sulphonal  and  Trional 191 

Santonin 191 

Constitution;  Behavior  in  the  organism;  Detection 
Sulphonal 193 

Preparation;  Detection;  In   urine;  Detection   of  hsematopor- 

phyrin  in  urine. 
Trional 196 

2.  Powerful   Organic   Substances   of   Rare    Occurrence    in  Toxi- 

coLOGicAL  Examinations 
Cantharidin 196 

Constitution;  Detection. 
Cytisine 198 

Preparation;  Toxic  action;  Detection. 
Digitalis  Bodies 200 

Digitonin,  Digi toxin,  Digitalinum  verum. 
Ergot 202 

Alkaloids;  Sclererythrin ;  Detection    of  ergot  in  flour;    Detec- 
tion and  estimation  of  the  alkaloids. 
Opium 205 

Meconicacid;  Meconin;  vSelenious-Sulphuric  acid,  a  reagent  for 

opium  alkaloids. 


CONTENTS  XIU 

Page 
Papaverine 208 

Constitution;  Detection. 

Pilocarpine 210 

Ptomaines 212 

Saponins ■ 213 

Physiological  action;  Detection  in  foaming  beverages,  such  as 

beer,   etc.;  Detection   of   githagin  in  flour. 

Haemolysis  and  Physiological  Salt  Solution 216 

Solanine 217 

Toxic  action;  Detection. 
Thebaine 220 

Constitution;  Detection. 

TOXALBUMINS 221 

Abrin,  Ricin;  Crotin;  Coagulation  of  blood  and  defibrinated 
blood. 

CHAPTER  V 

Special  Methods 

Quantitative  Estimation  of  Phosphorus  in  Phosphorated  Oils    224 
I.  W.  Straub's  method;  2.  A.  Frankel's  and  C.  Stich's  method. 

Special  Methods  for  Detecting  Arsenic 226 

Separation  of  arsenic  as  arsenic  trichloride 226 

Electrolytic  detection  of  arsenic 226 

Destruction  of  organic  matter  and  detection  of  arsenic  by  A. 

Gautier  and  G.  Lockemann 227 

Electrolytic  estimation  of  minute  quantities  of  arsenic  by    C. 

Mai  and  H.  Hurt 230 

Quantitative  estimation  of  arsenic  and  antimony  by  the  Gutzeit 

method 233 

Biological  detection  of  arsenic  by  means  of  penicillium  brevi- 

caule 235 

Detection  of  arsenic  in  organic  arsenic  compounds 238 

Cacodylic  acid;  Arrhenal;  Atoxyl;  In  urine; 

Quantitative  estimation  of  minute  quantities  of  arsenic  by  Karl 

Th.  Morner 240 

Detection  of  Salicylic  Acid  in  Foods  and  Beverages;  In  Wine, 

Meat  Products,  Milk 243 

Maltol 244 

Use  of  Chloral  Hydrate  in  Toxicological  Analysis  by  R.  Mauch. 

Alkaloidal  Estimations 244 

1.  By  the  picrolonate  method  of  H.  Matthes 246 

2.  By  precipitation  with  potassium  bismuthous  iodide  and  de- 
composition of  the  precipitate  with  alkali  hydroxide-carbonate 

by  H.  Thoms 248 

3.  By  the  method  of  H.  M.  Gordin 250 

Quantitative  estimation  of  strj^chnine  and  quinine  in  presence 

of  each  other 251 


xiv  CONTENTS 


Page 


Estimation  of  the  toxicity  of  chemical  compounds  by  blood 
haemolysis  by  A.  J.  J.  Vandevelde 251 

CHAPTER  VI 

Quantitative  Estimation  of  Alkaloids  and  other  Powerful  Sub- 
stances IN  Raw  Materials  and  in  their  Preparations 

Alkaloidal  Estimations  of  Drugs  and  Their  Pharmaceutical  Prepa- 
rations According  to  the  German  Pharmacopoeia.    .    .    .     253 

Estimation  of  alkaloid  in  aconite  root 254 

Estimation  of  cantharidin  in  Spanish  fly 256 

Estimation  of  cinchona  alkaloids 257 

I.  In  cinchona  bark ;  2.  In  aqueous  extract  of  cinchona  and 
in  alcoholic  extract  of  cinchona. 
Estimation  of  quinine  in  mixtures  of  cinchona  alkaloids  by  the 
sulphate  method 261 

I.  Cinchona  bark;  2.  Cinchona  extract. 
Estimation  of  colchicin  in  colchicum  seeds  and  in  colchicum 

corms 262 

Estimation  of  alkaloid  in  pomegranate  bark 264 

Estimation  of  caffeine  in  coffee,  tea,  kola  nuts  and  Guarana 
paste 264 

I.  C.  C.  Keller's  method;  2.  A.  Hilger-A.  Juckenack's 
method.  3.  A.  Hilger-H.  Gockel's  method;  4.  Socolof- 
Trillich-Gockel-method;  5.  E.  Katz's  method;  6.  K. 
Dieterich's  method. 

Estimation  of  alkaloid  in  ipecacuanha  root 270 

Estimation  of  nicotine  in  tobacco 272 

I.  R.   Kissling's method;   2.  C.   C.   Keller's  method;   3.    J. 
Toth's  method. 

Estimation  of  hydrastine  in  hydrastis  extract 274 

Estimation  of  berberine 275 

Estimation  of  hydrastine  by  the  picrolonate  method  of  H. 
Matthes  and  O.  Rammstedt 275 

I.  In  fluid  extract  of  hydrastis;  2.  In  hydrastis  root. 
Estimation  of  morphine  in  opium  and  in  its  pharmaceutical 
preparations 276 

I.  In  opium;  2.  In  extract  of  opium;  3.    In   wine  of  opium 
and  in  tincture  of  opium. 
Estimation  of  pilocarpine  in  jaborandum  leaves 279 

I.  G.  Promme's   method;  2.  H.    Matthes   and   O.    Ramm- 
stedt's   method. 
Piperine  and  its  estimation  in  pepper 281 

I.  J,  Konig's  method;  2.  Cazeneuve  and  Caillot's  method. 
Estimation  of  santonin  in  wormseed 282 

I.  K.  Thaeter's  method;  2.  J.  Katz's  method. 
Estimation  of  solanine  in  potatoes 284 


CONTENTS  XV 

Page 
I.  O.  Schmicdebcrg  and  G.  Meyer's  method;   i.  F.  v.  Mor- 
genstern's  method. 
Estimation  of  alkaloid  in  nux  vomica  and  its  preparations   .    .     286 

C.  C.  Keller's  method 286 

Method  of  the  German  Pharmacopoeia 287 

I.  In  nux  vomica;  2.  In  extract  of   nux   vomica;    3.  In 
tincture  of  nux  vomica. 

H.  Matthes  and  O.  Rammstedt's  method 289 

I.  In  extract  of  nux  vomica;  2.  In  tincture  of  nux  vomica; 
3,     In  nux  vomica. 
Estimation  of  strychnine  in  mixtures  of  strychnine  and  brucine 

by  C.  C.  Keller— H.  M.  Gordin 291 

Estimation  of  theobromine  and  cafifeine  in  cacao  and  in  choco- 
late    291 

Estimation  of  alkaloid  in  the  leaves  of  atropa  belladonna,  hyo- 

scyamus  niger  and  datura  strammonium 293 

Estimation  of  alkaloid  in  extract  of  belladonna,  according  to 

the  German  Pharmacopoeia,  in  extract  of  hyoscyamus   .    .    .     294 

Assay  of  officinal  extracts  by  E.  Merck 295 

Extract    of    belladonna;  Extract    of    cinchona;  Extract  of 
strychnine. 

CHAPTER  VII 

Detection  of  Carbon  Monoxide  Blood,  Blood  Stains  and  Human 

Blood 

1.  Recognition  of  carbon  monoxide  blood 297 

2.  Detection  of  blood  stains 300 

Haematin 301 

Spectroscopic  detection  of  blood 303 

Other  tests  for  blood 305 

Schonbein-van   Deen's   test;  Vitali's    procedure    in    this 
test;  Schaer's  procedure;  Aloin  test. 

3.  Biological  detection  of  human  blood 307 

APPENDIX 
Preparation  of  Reagents 

A.  General  alkaloidal  reagents 310 

B.  Special  reagents  and  solutions 313 

C.  The  indicator  iodeosine 315 

Index 317 


INTRODUCTION 


Nearly  all  the  common  poisons  and  drugs  may  be  placed  in 
one  of  three  groups.  This  classification,  based  upon  the 
chemical  behavior  of  these  substances  during  isolation  from  mix- 
tures is  as  follows: 

Group  I. — The  members  of  this  group  volatilize  without 
decomposition  when  heated  and  distil  from  an  acid  solution  with 
steam.  Yellow  phosphorus,  hydrocyanic  acid,  carbohc  acid, 
chloroform,  chloral  hydrate,  iodoform,  anihne,  nitrobenzene, 
carbon  disulphide  and  alcohol  are  the  principal  substances  of 
this  class. 

Group  II. — The  members  of  this  group  are  non-volatile, 
organic  substances  which  do  not  distil  from  an  acid  solution 
with  steam.  But  hot  alcohol  containing  tartaric  acid  will 
extract  them  from  extraneous  matter.  Alkaloids,  many  glu- 
cosides  and  bitter  principles,  as  well  as  certain  synthetic  organic 
drugs  like  acetanilide,  phenacetine,  antipyrine,  pyramidone, 
sulphonal  and  veronal  comprise  this  group. 

Group  III. — This  group  includes  all  poisonous  metals. 

In  toxicological  analysis,  therefore,  poisons  are  divided  into 
three  groups,  each  of  which  has  its  own  special  methods  of  pro- 
cedure. A  few  poisons  hke  mineral  acids,  caustic  alkalies, 
oxalic  acid  and  potassium  chlorate  cannot  be  conveniently 
placed  in  these  three  groups  owing  to  differences  in  solubihty 
and  other  peculiarities.  Special  tests  for  such  substances 
must  be  made  with  a  separate  portion  of  material.  Chapter  IV 
contains  a  description  of  the  methods  used  in  identifying  these 
substances. 

The  material  must  be  thoroughly  mixed  and  divided  into 
three  or  four  approximately  equal  portions,  unless  the  analysis 
is  to  be  hmited  to  the  detection  of  a  single  well-defined  substance. 
One  portion  is  tested  for  non- volatile,  organic  substances  (Chap- 

1 


Z  INTRODUCTION 

ter  II).  The  second  portion  is  examined  for  volatile  poisons 
(Chapter  I)  .and  the  residue  from  this  portion  is  used  in  testing 
for  poisonous  metals  (Chapter  III).  The  third  portion  is 
tested  for  substances  considered  in  Chapter  IV.  The  fourth 
portion  is  held  in  reserve  in  case  additional  material  is  needed 
to  verify  a  doubtful  result,  or  to  replace  a  portion  accidentally 
lost   during   analysis. 

Occasionally  it  is  advisable  to  depart  from  the  general  pro- 
cedure and  follow  a  special  method,  especially  in  detecting  a 
single  poison,  or  in  estimating  it  quantitatively.  For  instance, 
pure  ether  would  not  be  the  best  solvent  to  use  in  extracting 
strychnine  quantitatively  from  an  alkaline  solution.  A  mix- 
ture of  ether  and  chloroform,  or  better  pure  chloroform  would 
be  preferable,  since  strychnine  is  more  soluble  in  the  latter 
solvent  than  in  pure  ether.  For  the  same  reason  chloroform 
should  be  used  in  the  quantitative  extraction  of  caffeine  or 
antipyrine.  When  only  a  small  quantity  of  material  is  available 
for  analysis,  tests  for  all  three  groups  of  poisons  may  be  made 
with  the  same  portion.  In  this  case  after  removal  of  volatile 
poisons  (Chapter  I)  the  residue  should  be  divided  into  two  un- 
equal portions.  The  larger  portion  should  be  tested  for  non- 
volatile, organic  poisons  (Chapter  II).  The  smaller  portion 
together  with  the  residue  left  after  extracting  non-volatile,  or- 
ganic poisons  should  be  tested  for  poisonous  metals  (Chapter 
III).  It  is  advisable,  however,  even  in  such  a  case  to  reserve  a 
portion  of  material  for  any  contingencies. 

Organs  of  the  human  body  like  liver,  kidneys,  spleen,  heart, 
brain,  stomach  or  intestines  with  contents  should  be  cut  into 
small  pieces  and  then  finely  chopped  before  being  examined 
chemically.  An  organ  should  first  be  cut  into  small  pieces  with 
sharp,  clean  scissors  and  then  minced  with  a  clean  chopping 
knife  in  a  new  wooden  bowl,  or  a  small  meat  machine,  which 
has  been  carefully  cleaned,  may  be  used.  Material  may  be 
held  with  nickel  plated  tongs  while  being  cut  with  scissors. 


DETECTION  OF  POISONS 


CHAPTER  I 
VOLATILE  POISONS 


Yellow  Phosphorus  and  Other  Poisons  Volatile  from  Acid  Solution 

with  Steam 

Scherer's  Test. — This  test  should  precede  the  distillation 
described  on  page  i8.  The  principle  of  the  test  is  that  moist 
phosphorus  vapor  and  silver  nitrate  form  black  silver  phosphide 
(AgsP),  metallic  silver,  phosphoric  and  sometimes  phosphorous 
acid.  Place  the  finely  divided  material  in  a  small  flask  and 
cover  with  water  if  a  sufficient  quantity  is 
not  present.  Cut  a  V-shaped  slit  in  the 
cork  and  place  the  latter  loosely  in  the 
mouth  of  the  flask  so  that  the  two  strips  of 
filter  paper  are  freely  suspended  (Fig.  i). 
Moisten  one  strip  with  silver  nitrate  and 
the  other  with  lead  acetate  solution.^ 
Warm  gently  upon  the  water-bath  (40  to 
50°).^  If  the  silver  paper  is  darkened  but 
not  the  lead  paper,  yellow  phosphorus  njay 
be  present.  If  both  papers  are  darkened, 
hydrogen  sulphide  also  is  present.  In  the 
latter  case  yellow  phosphorus  may  be  pres- 
ent with  hydrogen  sulphide.  In  absence  of 
hydrogen  sulphide,  darkening  of  the  silver 
paper  is  not  final  proof  of  yellow  phosphorus,  for  any  volatile 
organic  substance  having  reducing  properties,    as   formalde- 

1  A  more  sensitive  "lead  paper"  may  be  obtained  by  using  alkaline  lead  solu- 
tion prepared  by  adding  excess  of  sodium  hydroxide  to  the  solution  of  a  lead 
salt  whereby  Pb(OH)(ONa)  and  Pb(0Na)2  are  formed. 

^  Temperatures  in  this  book  are  expressed  in  Centigrade  degrees.     Tr. 

3 


Fig. 


4  DETECTION   OF   POISONS 

hyde    (H.CHO),   or  formic   acid    (H.COOH),    may  give  the 
same  result.' 

Scherer's  test  is  of  value  in  proving  the  absence  rather  than  the  presence  of 
yellow  phosphorus.  It  is  a  good  preliminary  test,  as  it  excludes  phosphorus  if 
the  silver  paper  is  unchanged. 

Distillation. — Place  a  portion  of  finely  divided  and  thoroughly 
mixed  material  in  a  large  round-bottom  flask  and  add  enough 
distilled  water  for  free  distillation.  Then  add  tartaric  acid 
solution  drop  by  drop  until  the  mixture  is  acid  after  thorough 
shaking.  Practice  analyses^  usually  require  20  to  30  drops  of 
10  per  cent,  tartaric  acid  solution. 

In  examining  animal  material,  as  the  stomach  or  intestines 
and  contents,  or  organs,  like  liver,  spleen  and  kidneys,  it  is  often 
unnecessary  to  add  much  water  because  enough  is  usually  pres- 
ent. First  chop  the  material  in  a  wooden  tray  with  a  steel  knife. 
In  a  medico-legal  analysis  the  tray  should  be  new.  A  meat 
machine  which  has  been  carefully  cleaned  may  be  used.  Thin 
the  material  with  a  little  distilled  water,  acidify  with  dilute 
tartaric  or  sulphuric  acid  and  finally  distil. 

If  Scherer's  test  is  positive,  begin  distilling  with  the  Mit- 
scherlich  apparatus  (Fig.  2) ;  but  if  negative,  distil  in  the  usual 
way  with  the  Liebig  condenser  (see  page  18).  The  distillate 
may  contain: 

Yellow  phosphorus  Nitrobenzene 

Hydrocyanic  acid  Aniline 

Carbolic  acid  Ethyl  alcohol 

Chloroform  Acetone 

Chloral  hydrate  Carbon  disulphide 

Iodoform  Benzaldehyde 
Bitter  almond  water 

'Laboratory  practice  in  detecting  poisons  may  be  given  by  mixing  small  quan- 
tities (from  0.03  to  0.05  or  o.i  gram)  of  a  poison  with  dry  bread  or  biscuit 
crumbs,  meal  or  meat.  Finely  chopped  organs  (liver,  kidney,  spleen,  etc.), 
sausage  meat,  beer,  wine  or  milk  may  be  used.  Drugs  like  morphine,  codeine, 
qiunine,  acetanilide,  phenacetine,  antipyrine,  caffeine,  santonin,  sulphonal, 
veronal,  calomel,  tartar  emetic,  subnitrate  of  bismuth,  etc.,  may  be  mixed  with 
powdered  cane-  or  milk-sugar.  The  last  kind  of  practice  is  especially  suitable 
for  students  of  pharmacy. 


VOLATILE   POISONS 
YELLOW  PHOSPHORUS 


Mitscherlich  Method  of  Detecting  Yellow  Phosphorus 

The  principle  of  this  method  is  that  yellow  phosphorus  is 
volatile  with  steam  and  becomes  luminous  in  contact  with  air. 
The  phosphorescence  is  best  seen  in  a  dark  room. 


Fig.  2. — Mitscherlich  Apparatus. 

Procedure. — Arrange  the  apparatus  as  in  Fig.  2.  Support 
the  condenser  in  a  vertical  position  and  connect  the  upper  end 
with  the  flask  by  a  glass  tube  about  8  mm.  internal  diameter. 
This  tube  has  two  right-angle  bends  and  each  end  passes  through 


b  DETECTION   OP   POISONS 

a  cork.  Have  condenser  and  tube  scrupulously  clean  to  avoid 
interference  -with  the  phosphorescence. 

Have  the  flask  at  most  not  more  than  a  third  full.  This  pre- 
caution is  necessary  because  many  materials,  containing  protein 
substances  like  albumin,  albumose,  etc.,  and  starchy  matter, 
when  distilled  in  aqueous  solution,  cause  more  or  less  foaming 
which  is  liable  to  carry  over  solid  matter  into  the  receiver.  Use 
as  the  receiver  an  Erlenmeyer  flask  containing  a  little  distilled 
water  (3  to  5  cc.)  into  which  the  end  of  the  condenser  dips. 
This  precaution  prevents  loss  of  easily  volatile  substances  Hke 
hydrocyanic  acid  and  chloroform.  Heat  the  flask  upon  a  wire 
gauze  of  fine  mesh,  asbestos  plate  or  sand  bath  and  bring  the 
contents  to  boiling  by  raising  the  temperature  gradually. 
There  is  some  danger  of  burning  or  carbonizing  organic  matter 
on  the  bottom  of  the  flask,  if  heat  is  applied  too  strongly  or 
rapidly.  When  boiling  begins,  make  the  room  as  dark  as  pos- 
sible and  watch  for  phosphorescence  in  the  tube  and  condenser. 
It  usually  appears  as  a  luminous  ring  or  band  in  the  upper  part 
of  the  condenser.  When  this  is  distinctly  visible,  the  presence 
of  yellow  phosphorus  is  established.  Phosphorescence  during 
distillation  with  steam  is  very  characteristic  of  yellow  phos- 
phorus and  frequently  is  the  only  sure  and  unquestionable  test 
for  this  element. 

Phosphorescence  is  a  process  of  oxidation  by  which  phos- 
phorus vapor  is  changed  to  phosphorous  acid.  Should  it  not 
appear  immediately,  distillation  must  be  continued  for  some 
time,  since  certain  substances  hke  ethyl  alcohol,  ether,  turpen- 
tine and  many  other  ethereal  oils  either  prevent  the  phenomenon 
entirely  or  seriously  retard  it.  Considerable  carbolic  acid, 
creosote,  chloroform,  chloral  hydrate,  as  well  as  hydrogen 
sulphide,  may  completely  prevent  phosphorescence, 

K.  Polstorff  and  J.  Mensching^  have  shown  that  mercuric 
chloride  as  well  as  other  mercury  compounds  may  also  interfere 
with  phosphorescence.  Possibly  mercuric  chloride  carried  over 
by  steam  is  reduced  to  metallic  mercury  by  phosphorus  vapor. 
In  that  case  the  metal  should  appear  in  the  distillate.     The 

^Berichte  der  Deutschen  chemischen  Gesellschaft  19,  1763  (iJ 


VOLATILE   POISONS 


fact  that  both  metallic  mercury  and  phosphoric  acid  can  be 
detected  in  the  distillate  favors  the  supposition  that  action 
takes  place  between  phosphorus  vapor  and  mercuric  chloride. 

Phosphorescence,  however,  often  appears  when  these  sub- 
stances have  passed  over.  But  even  when  prolonged  dis- 
tillation fails  to  give  a  positive  result,  this  must  not  be  accepted 
as  final  proof  of  the  absence  of  phosphorus  until  other  tests 
have  been  made.  Whatever  the  result,  evaporate  a  portion 
of  the  distillate  to  dryness  on  the  water-bath  with  excess  of 
saturated  chlorine  water,  or  with  a  little  fuming  nitric  acid. 
Phosphorus  always  imparts  a  strong  odor  to  the  distillate. 
Small  drops  of  phosphorus  appear  if  the  quantity  is  large,  and 
the  solution  contains  phosphorous  acid.  Dissolve  the  residue 
from  evaporation  in  2  to  3  cc.  of  water  and  test  in  two  sepa- 
rate portions  for  phosphoric  acid. 

1.  Ammonium  Molybdate  Test. — Acidify  the  solution  with 
a  few  drops  of  concentrated  nitric  acid.  Add  an  equal  volume 
of  ammonium  molybdate  solution 
and  warm  to  about  40°.  Phos- 
phoric acid  precipitates  yellow 
ammonium  phospho-molybdate. 

2.  Ammonium  Magnesium 
Phosphate  Test. — Add  magnesia 
mixture^  to  the  second  portion. 
Phosphoric  acid  gives  a  white 
crystalline  precipitate  of  ammo- 
nium magnesium  phosphate  (H4N) 
MgP04.6H20.  Vigorous  shaking 
favors  precipitation.  When  only 
traces  of  phosphoric  acid  are  present,  long  standing  is  necessary 
before  the  precipitate  appears.  Always  examine  the  precipi- 
tate with  the  microscope.     It  should  consist  of  well-formed 


Fig.  3. — Ammonium  magnesium 
phosphate  crj-stals. 


^Magnesia  mixture  is  a  clear  solution  prepared  by  mixing  equal  volumes  of 
magnesium  chloride,  ammonium  chloride  and  ammonium  hydroxide  (about 
10  per  cent.)  solutions.  It  contains  the  readil}'  soluble  double  chloride  of  ammo- 
nium and  magnesium  which  is  not  decomposed  by  ammonium  hydroxide.  This 
reagent  is  prepared  as  needed  and  should  be  perfectly  clear  and  colorless. 


8  DETECTION   OF   POISONS 

crystals  or  at  least  should  be  crystalline.      These  crystals  are 
transparent,  acicular  prisms  (Fig.  3). 

Notes. — A.  Fischer^  states  that  substances  interfering  more  or  less  with  the 
detection  of  phosphorus  by  the  Mitscherlich  method  are  usually  less  troublesome 
if  Hilger  and  Nattermann's  procedure  is  used  (see  page  15).  The  essential 
feature  of  the  latter  process  consists  in  allowing  steam  charged  with  phosphorus 
to  pass  into  the  air,  or  in  admitting  air  into  the  apparatus. 

Detection  of  Phosphorus  and  Phosphorous  Acid 

(Blondlot^-Dusart^) 

When  the  Mitscherlich  method  fails  to  show  phosphorus,  it 
is  often  necessary  to  test  for  phosphorous  acid.  This  is  the 
first  product  in  the  oxidation  of  phosphorus  and  is  easily  formed. 
The  Blondlot-Dusart  method  shows  the  slightest  trace  of  phos- 
phorous (H3PO3)  and  hypophosphorous  (H3PO2)  acids  as  well 
as  yellow  phosphorus.  The  method  consists  in  converting 
yellow  phosphorus  into  phosphine  (PH3)  by  nascent  hydrogen. 
The  lower  oxidation  products  of  phosphorus,  namely,  hypo- 
phosphorous  (H3PO2)  and  phosphorous  (H3PO3)  acids, ^  when 
warmed  with  zinc  and  dilute  sulphuric  acid  are  reduced  to 
phosphine  by  nascent  hydrogen: 

H3PO2  +  4H  =  PH3  +  2H2O, 

H3PO3  +  6H  =  PHs  +  3H2O. 

Phosphine,   or  hydrogen   charged  with  phosphorus  vapor, 
burns  with  a  characteristic  green  flame  (Dusart's  reaction) : 
2PH3  +  4O2  =  P2O5  +  3H2O. 

The  green  flame  is  easily  recognized  by  depressing  a  cold 
porcelain  dish  or  plate  upon  the  flame.  Detection  of  phosphorus 
by  the  Blondlot-Dusart  method  depends  upon  these  two  facts. 

A  toxicological  analysis  usually  deals  with  the  detection  of 
traces  of  yellow  phosphorus.  Hydrogen  after  acting  in  the 
nascent  state  upon  the  material  is  not  directly  examined  for 

^  Pfiueger's  Archiv  97,  578  (1903). 

2  Journal  de  Pharmacie  et  de  Chimie  (3),  41,  25. 

2  Comptes  rendus  de  I'Academie  des  Sciences,  43,  11 26. 

^  Nascent  hydrogen  will  not  reduce  ordinary,  or  ortho-phosphoric  acid  (H3PO4), 
and  its  derivatives,  pyrophosphoric  (H4P2O7)  and  meta-phosphoric  (HPO3)  acids, 
to  phosphine. 


VOLATTTJC   rOIRONS  « 

phosphorus  but  is  first  passed  into  dilute  silver  nitrate  solution. 
Phosphorus  and  phosphine  precipitate  black  silver  phosphide^ 

(AgaP): 

PH3  +  sAgNOa  =  Ag3P  +  3HNO3. 

Thus  traces  of  yellow  phosphorus  may  be  concentrated  in  the 

silver  precipitate  from  which  nascent  hydrogen  will  liberate 

phosphine: 

Ag:,P  +  3H  =  PH3  +  sAg. 

If  hydrogen  produces  a  black  or  gray  precipitate  in  the  silver 
solution,  phosphorus  is  not  necessarily  present,  as  hydrogen 
sulphide,  arsine,  stibine  and  reducing  organic  compounds  be- 
have similarly  with  silver  nitrate.  A  black  precipitate  there- 
fore should  always  be  examined  for  phosphorus  by  the  Dusart 
reaction. 

In  the  detection  of  yellow  phosphorus,  the  Blondlot-Dusart 
method  combines  two  distinct  operations,  namely: 

1.  Preparation  of  the  silver  phosphide  precipitate. 

2.  Examination  of  this  precipitate  in  the  Dusart  apparatus. 
Procedure,     i.  Preparation  of  Silver  Phosphide. — Thin  the 

finely  divided  material  with  water  in  a  capacious  flask  where 
hydrogen  is  being  evolved  from  phosphorus-free  zinc  and  pure 
dilute  sulphuric  acid  (1:5).  In  testing  for  phosphorous  acid 
alone  (see  page  14)  use  the  filtrate  from  an  aqueous  extract 
of  the  material,  or  the  filtrate  from  the  residue  left  after  the 
MitscherKch  distillation  (see  page  5).  Nascent  hydrogen 
should  act  for  1.5  to  2  hours,  or  even  longer,  and  pass  through 
neutral  silver  nitrate  solution  in  the  receiver  at  the  end  of  the 
apparatus.  If  yellow  phosphorus  is  present,  the  hydrogen  will 
contain  phosphorus  and  phosphine  and  cause  a  black  precipi- 
tate of  silver  phosphide  in  the  silver  nitrate  solution.  Collect 
the  precipitate  upon  a  small  ash-free  paper,  wash  with  a  Httle 
cold  water  and  examine  in  the  Dusart  apparatus  as  described 
elsewhere. 

1  Phosphorus  cannot  be  determined  quantitatively  as  silver  phosphide  because 
this  compound  is  partially  decomposed  by  water.  Phosphoric  and  phosphorous 
acids  pass  into  solution: 

(a)  2Ag3P  +  5O  +  3H2O  =  6Ag  +  2H3PO4, 

(b)  2Ag3P  +  3O  +  3H2O  =  6Ag  +  2H3PO3. 


10 


DETECTION    OF   POISONS 


If  there  is  silver  phosphide  in  the  precipitate,  the  filtrate  will 
contain  phosphoric  or  phosphorous  acid  (see  Note,  page  9). 
To  detect  phosphoric  acid,  first  add  hydrochloric  acid  to  remove 
excess  of  silver  from  this  filtrate.  Filter  through  paper  pre- 
viously well  washed  with  acid  and  water  and  completely  expel 
hydrochloric  acid  from  the  filtrate  by  evaporation  upon  the 
water  bath  with  concentrated  nitric  acid.  Dissolve  the  resi- 
due in  a  little  warm  water  and  test  for  phosphoric  acid  with  am- 
monium molybdate  or  magnesia  mixture. 

2.  Examination  of  the  Silver  Precipitate  (AgsP)  for  Phos- 
phorus.— Two  forms  of  apparatus  may  be  used  for  this  purpose, 
namely : 

(a)  Fresenius-Neubauer^  Apparatus. — Generate  hydrogen 
in  flask  A  (Fig.  4)  from  pure  phosphorus-free  zinc  and  dilute 


Fig.  4. — Fresenius-Neubauer  Apparatus. 


sulphuric  acid.  Fill  U-tube  C  with  pieces  of  pumice  stone  satu- 
rated with  concentrated  potassium  hydroxide  solution  to  ab- 
sorb any  hydrogen  sulphide.     Use  hard  glass  for  tube  D  and 

^  C.  R.  Fresenius,  Qualitative  chemische  Analyse,  XVI  edition,  page  521. 


VOLATILE  POISONS  1 1 

have  the  tip  F  of  platinum.  The  part  marked  E^  is  a  glass 
stop-cock  or  screw-tap.  Reservoir  B  serves  to  hold  liquid  from 
A  when  cock  E  is  closed.  A  platinum  tip  is  essential,  other- 
wise the  flame  instead  of  being  colorless  will  always  be  yellow 
from  sodium  in  the  glass.  The  place  where  the  platinum  tip  is 
fused  into  the  glass  should  be  cooled  by  wrapping  cotton  around 
the  glass  and  keeping  it  moist. 

Procedure. — Open  E  and  let  hydrogen  from  A  pass  for  some 
time  through  the  apparatus  to  expel  air.  Then  close  E  and 
liquid  in  A  will  rise  into  B.  Now  open  E  just  enough  to  allow 
hydrogen  to  burn  with  a  small  flame  which  should  be  colorless 
in  the  dark.  If  there  is  no  trace  of  green  in  the  inner  cone  and  a 
porcelain  dish  depressed  upon  the  flame  does  not  show  an  em- 
erald green  coloration,  hydrogen  is  phosphorus-free.  It  is  well 
to  repeat  this  test.  To  test  the  silver  precipitate  for  phosphorus, 
wash  it  with  the  paper  into  B  with  a  little  water. 

When  the  entire  precipitate  is  in  A,  close  E  until  all  the  liquid 
has  risen  from  A  into  B.  Then  open  E,  Hght  the  hydrogen  and 
examine  the  flame  in  the  dark.  If  the  precipitate  contains  a 
trace  of  silver  phosphide,  the  inner  cone  will  b*e  green  and  a 
porcelain  dish  depressed  upon  the  flame  will  show  an  emerald 
green  coloration.  Have  the  hydrogen  flame  small  so  that  its 
color  may  be  observed  for  some  time. 

(b)  Hilger-Nattermann^  Apparatus. — Reduction  takes  place 
in  a  loo  cc.  flask  closed  by  a  rubber  stopper  with  three  holes, 
two  of  which  are  for  right- angle  tubes  just  passing  through  the 
stopper  and  the  third  for  a  thistle  tube  going  to  the  bottom  of 
the  flask  (Fig.  5).  Hydrogen  from  a  Kipp  generator  enters  the 
flask  by  one  tube  and  leaves  by  the  other.  Attach  to  the  latter 
a  U-tube  filled  with  pieces  of  pumice  stone  saturated  with 
concentrated  potassium  hydroxide  solution  to  absorb  hydrogen 
sulphide.     Connect  the  other  end  of  the  U-tube  with  a  hard 

^  Fresenius  and  Neubauer  use  a  screw  pinch-cock  instead  of  a  gas-cock  but  by 
means  of  a  short  rubber  connector  they  interpose  an  ordinarj'  cock  between  the 
gas  flask  A  and  the  U-tube  C. 

2  Forschungsbericht  iiber  Lebensmittel  und  ihre  Beziehungen  zur  Hygiene, 
etc.,  4,  241-258  (1897). 


12 


DETECTION   OF   POISONS 


glass  tube  tipped  with  platinum.^  Cut  the  paper  containing 
the  precipitate  into  small  pieces  and  place  in  the  flask  which 
contains  in  addition  a  few  pieces  of  phosphorus-free  zinc  and 
enough  water  to  seal  the  thistle  tube.  Light  the  hydrogen  after 
it  has  passed  through  the  apparatus  for  some  time  and  been 
found  free  from  air  by  the  usual  test.  Seen  in  the  dark  the 
flame  should  be  entirely  colorless  and  burn  without  a  green  cone 


Fig.  5. — -Hilger-Natterman  Apparatus. 

• 

or  a  greenish  glow.^  Hilger  and  Nattermann  advise  a  spectro- 
scopic examination  of  the  flame  to  determine  the  purity  of  the 
zinc.  Pure  zinc  gives  a  hydrogen  spectrum  which  shows  only 
an  orange  colored  line  in  place  of  the  yellow  sodium  line.  The 
minutest  trace  of  phosphorus  will  give  three  green  lines  lying 
to  the  right  of  the  line  D.  The  color  of  two  of  these  lines  is  more 
pronounced  than  that  of  the  third.  Having  thus  tested  the 
purity  of  zinc  and  sulphuric  acid,  pour  a  few  cc.  of  dilute  sul- 
phuric acid  (i  :  5)  through  the  thistle  tube  into  the  flask  con- 
taining zinc  and  the  silver  precipitate.  If  the  latter  contains 
phosphorus,  the  flame  will  show,  though  not  always  at  once,  a 
green  coloration  which  should  be  examined  with  the  spectro- 
scope. 

^  Hilger  and  Nattermann  use  a  platinum  tipped  blow-pipe  instead  of  a  glass 
tube  tipped  with  the  same  metal.  Cotton,  which  is  kept  moist  and  acts  as 
a  cooler,  is  wrapped  around  the  blow-pipe  below  the  tip. 

2  Zinc  entirely  free  from  phosphorus  which  will  stand  this  test  is  difficult 
to  obtain. 


VOLATILE    POISONS  13 

The  Mitscherlich  method  affords  a  distillate  especially  suit- 
able for  the  Blondlot-Dusart  test.  If  this  imparts  a  green  color 
to  the  hydrogen  flame,  there  can  be  no  question  about  the  pres- 
ence of  phosphorus. 

Although  the  Blondlot-Dusart  test  is  very  delicate,  many 
chemists  refuse  to  accept  it  as  a  substitute  for  the  Mitscherlich 
test.  Selmi  states  that  animal  material  like  brain,  which  con- 
tains organic  phosphorus  compounds,  yields  after  putrefaction 
a  distillate  that  often  gives  a  black  precipitate  with  silver  ni- 
trate solution.  This  will  impart  a  green  tinge  to  the  hydrogen 
flame  in  the  Blondlot-Dusart  test. 


Z.  Haldsz,^  however,  has  failed  to  confirm  Selmi's  results.  He  examined  two 
kinds  of  animal  material  by  the  Blondlot-Dusart  method.  First,  he  tested  nor- 
mal brains  (man,  calf,  hog) ;  second,  the  brain  and  other  organs  of  rabbits  that 
had  been  given  poisonous  doses  of  phosphorus  by  the  mouth  and  subcutaneously. 
He  examined  these  organs  when  fresh  and  also  from  we6k  to  week  after  more  or 
less  pronounced  putrefaction  had  set  in,  but  could  not  detect  phosphorus  in  the 
brain  in  a  single  instance.  These  experiments  disprove  the  earlier  idea  that 
phosphorus  normally  present  in  the  brain  is  so  changed  during  putrefaction  that 
it  can  be  detected  by  the  Blondlot-Dusart  reaction.  He  also  failed  to  detect 
phosphorus  in  the  brain  of  rabbits  poisoned  by  this  element,  though  he  found  it  in 
other  organs,  as  stomach  and  intestines,  and  in  those  rich  in  blood,  as  liver, 
lungs  and  kidneys.  He  could  always  detect  small  or  large  quantities  of  phos- 
phorus in  any  organ  which  this  element  had  directly  reached,  or  by  which  it  had 
been  indirectly  absorbed.  If  any  compound  containing  phosphorus  is  really 
formed  in  the  brain  during  putrefaction,  Halasz  concluded  that  it  is  not  volatile 
with  steam  and  does  not  give  the  Blondlot-Dusart  reaction.  On  the  basis  of  these 
experiments  Halasz  holds  that  the  Blondlot-Dusart  method  of  detecting  phos- 
phorus is  just  as  reliable  for  forensic  purposes  as  that  of  Mitscherlich. 

Procedure  of  Halasz  in  the  Blondlot-Dusart  Method 

Make  a  thin  mixture  of  very  finely  divided  material  and  boUed  water  in  a  flat- 
bottom  flask  where  hydrogen  is  being  generated  from  phosphorus-free  zinc  and 
dilute  sulphuric  acid.  Warm  upon  the  water-bath  and  pass  the  gas  through  an 
absorption  tube  provided  with  several  bulbs  and  containing  neutral  silver  nitrate 
solution.  Concentrated  sulphuric  acid  and  a  little  platinic  chloride  may  be  added 
toward  the  end  to  hasten  the  evolution  of  gas.  Nascent  hydrogen  thus  acts  upon 
phosphorus  in  the  animal  material  for  2-2.5  hours.  Finallj'  wash  the  silver 
precipitate  carefulh'  with  water  and  transfer  it  with  the  paper  to  the  Blondlot- 
Dusart  apparatus. 

^Zeitschrift  fiir  anorganische  Chemie  26,  43S  (1901). 


14  DETECTION   OE   POISONS 

Detection  of  Phosphorous  Acid 

The  reduction  of  phosphorous  acid  to  phosphine  by  zinc  and  dilute  siilphuric  acid 
takes  place  very  slowly.  Hilger  and  Nattermann  state  that  even  a  few  milU- 
grams  require  the  action  of  nascent  hydrogen  for  lo  to  14  days.  Moreover  care- 
ful manipulation  is  necessary  because  silver  phosphide  is  quite  unstable.  Water 
decomposes  this  substance  into  metallic  silver  and  phosphorous  acid  and  the 
nitric  acid  present  oxidizes  the  latter  to  phosphoric  acid.  Therefore  when  special 
attention  must  be  given  to  phosphorous  acid,  Hilger  and  Nattermann  recommend 
examining  the  silver  precipitate  (presumably  AgaP)  after  2  days,  or  at  most 
3,  for  phosphorus  by  the  Blondlot-Dusart  method  and  the  filtrate  for  phos- 
phoric acid  (see  page  10). 

Detection  of  Phosphorus  in  Phosphorated  Oils 

1.  Straub's^  Test. — If  a  phosphorated  oil  is  placed  on  the 
surface  of  copper  sulphate  solution,  phosphorus  will  gradually 
pass  from  the  oil  to  the  aqueous  solution  and  first  form  black 
copper  phosphide.  The  latter,  acting  as  a  carrier  of  oxygen, 
oxidizes  phosphorus  still  in  the  oil  to  phosphoric  acid  which 
dissolves  in  the  water. 

Shake  lo  cc.  of  phosphorated  oil  in  a  test-tube  with  5  cc.  of  i 
per  cent,  copper  sulphate  solution.  According  to  the  amount  of 
phosphorus  a  black  or  light  brown  coloration  will  appear  in  the 
emulsion  at  once,  or  in  a  few  minutes,  and  at  most  after  2 
hours.  Phosphoric  acid  in  the  aqueous  solution  may  be  recog- 
nized by  ammonium  molybdate.  At  least  0.0025  P^^  cent,  of 
phosphorus  may  be  detected  in  this  way. 

A  practical,  therapeutic  application  of  this  reaction  may  be  made  in  acute  phos- 
phorus poisoning.  Administration  of  copper  sulphate  solution  may  prevent 
absorption  of  free  phosphorus  still  in  the  gastro-intestinal  tract. 

2.  The  Mitscherlich  test  is  also  apphcable  to  phosphorated 
oils,  even  though  the  oil  may  contain  only  0.0002  gram  of 
phosphorus  in  100  grams.  But  phosphorescence  will  not 
appear  unless  air  is  admitted  into  the  tube  from  time  to  time. 

Phosphorus  in  oils  cannot  be  determined  quantitatively  by 
the  distillation  method,  for  not  more  than  36  to  41  per  cent,  of 

^W.  Straub,  Miinchener  medizinische  Wochenschrift  50,1145;  Archiv  der 
Pharmazie  241,  335  (1903);  and  Zeitschrift  fiir  anorganische  Chemie  35,  460 
(1903)- 


VOLATILE   POISONS 


16 


phosphorus  will  distil  over.     The  quantitative  method  recom- 
mended by  Straub  (see  page  224)  may  be  used  in  that  case. 

Detection    and    Quantitative    Estimation    of    Phosphorus 
(Mitscherlich-Scherer) 

Acidify  a  weighed  portion  of  material  with  dilute  sulphuric 
acid  and  add  a  little  ferrous  sulphate.  Distil  in  a  gentle  stream 
of  carbon  dioxide,  using  a  large  flask  fitted  with  a  two-hole 
stopper.  Expel  air  completely  from  the  apparatus  by  carbon 
dioxide  before  heating.     Use  as  receiver  a  flask  fitted  with  a 


Fig,  6. — Hilger-Nattermann  Apparatus  for  Detecting  and  Quantitatively  Esti- 
mating Phosphorus. 

two-hole  Stopper.  Pass  the  end  of  the  condenser  through  one 
hole  and  connect  the  other  with  a  U-tube  containing  silver 
nitrate  solution.  Evaporate  the  distillate  upon  the  water 
bath  with  strong  bromine  water,  or  with  concentrated  nitric 
acid,  to  oxidize  phosphorus  or  any  phosphorous  acid  formed. 
Dissolve  the  residue  in  a  Httle  water  and  precipitate  phos- 


16  DETECTION   OF   POISONS 

phoric  add  with  magnesia  mixture.  Weigh  the  precipitate  as 
magnesium  pyrophosphate,  Mg2P207.  Heat  the  contents  of 
the  U-tube  with  concentrated  nitric  acid.  Precipitate  silver 
as  silver  chloride  and  filter.  Concentrate  the  filtrate  by  evapo- 
ration and  precipitate  phosphoric  acid  with  magnesia  mixture 
as  before.  Combine  this  precipitate  with  the  other.  In  dis- 
tillation some  phosphorus  separates  as  globules  in  the  first 
receiver  and  any  carried  over  as  vapor  by  carbon  dioxide  is  re- 
tained by  silver  nitrate  solution.  As  the  steam  distillation  of 
phosphorus  is  very  slow,  the  process  should  be  carried  on  for  at 
least  3  hours.  Hilger  and  Nattermann  recommend  the 
apparatus  in  Fig.  6  not  only  for  detecting  phosphorus  but  for 
estimating  it  quantitatively.  Air  may  be  mixed  with  phos- 
phorus vapor  by  means  of  stop-cock  K  and  the  characteristic 
phosphorescence  will  appear. 

Remarks  Upon  the  Mitscherlich  Test. — Hilger  and  Nattermann  state  that 
o .  00006  gram  of  yellow  phosphorus  is  the  smallest  quantity  that  can  be  detected 
by  the  Mitscherlich  method.  When  200  cc.  of  water  containing  o .  0003  gram  of 
phosphorus  were  distilled,  there  was  brilliant  phosphorescence  for  5  minutes. 
The  degree  of  dilution  seems  to  have  no  effect  upon  the  result,  at  least  not  within 
limits  occurring  in  practice.  Hydrogen  sulphide,  always  present  in  putrefying 
animal  matter,  has  no  apparent  effect  upon  phosphorescence.  Free  phosphorus 
can  be  detected  in  putrid  organic  matter  even  after  the  lapse  of  considerable  time. 
Putrefactive  and  digestive  processes  appear  to  prevent  oxidation  of  phosphorus. 
Dragendorff  detected  phosphorus  in  an  exhumed  body  several  weeks  after  death. 
Neumann  found  free  phosphorus  in  a  human  body  fourteen  days  and  Elvers 
eight  weeks  after  death. 

When  an  acid  aqueous  solution  is  distilled  in  the  Mitscherlich  apparatus,  the 
flask  residue  always  contains  phosphoric  (H3PO4),  phosphorous  (H3PO3)  and 
hypophosphorous  (H3PO2)  acids  and  red  phosphorus.  Distillation  of  a  solution 
of  0.0644  gram  of  phosphorus  gave  only  71.33  per  cent,  in  the  distillate.  The 
residue  contained: 

Phosphorus  as  phosphoric  acid             (H3PO4)  18.93  per  cent. 

Phosphorus  as  phosphorous  acid          (H3PO3)  2.15  per  cent. 

Phosphorus  as  hjrpophosphorous  acid  (H3PO2)  4.27  per  cent. 

Phosphorus  as  red  phosphorus  2 .  98  per  cent. 

28.33 

Oxidation  of  phosphorus  may  be  prevented  by  distilling  in  a  current  of  carbon 
dioxide  as  in  the  Mitscherlich-Scherer  method  (see  page  15). 

Metabolism  in  Phosphorus  Poisoning. — Phosphorus  has  a  very  poisonous 
action  upon  the  processes  of  metabolism.  Present  as  a  vapor  in  the  blood  and 
tissue  fluids,  it  retards  normal  oxidative  processes  occurring  in  the  animal  organ- 


VOLATILE   POISONS  17 

ism  during  metabolism.  In  pliosphorus  poisoning  the  usual  course  of  chemical 
metabolism  is  wholly  changed.  Fat  instead  of  being  oxidized  is  deposited  in  the 
organs  in  large  quantity  (fatty  degeneration  of  the  liver).  DifTerent  observers 
believe  there  is  formation  of  fat  from  protein.  During  phosphorus  poisoning  the 
quantity  of  protein  broken  down  is  greatly  increased.  In  human  metabolism 
this  applies  to  protein  in  both  food  and  tissues.  Yet  the  needs  of  the  organism 
are  not  satisfied  and  the  conclusion  is  that  the  changes  are  not  as  complete  as  in 
normal  protein  metabolism.  This  increase  in  the  breaking  down  of  tissues  in 
pliosphorus  poisoning  recalls  similar  changes  which  take  place  during  respiration 
in  insuflicient  oxygen.  Accompanying  these  abnormal  processes  arc  certain 
nitrogenous  and  non-nitrogenous  products  of  metabolism  which  either  are  not 
normally  formed  in  the  organism  or  appear  merely  as  intermediate  steps  in  the 
formation  of  the  oxidative  products  of  metabolism.  Decomposition  of  the  protein 
molecule  goes  in  part  only  as  far  as  the  amino  acids.  Consequently  in  phosphorus 
poisoning  the  urine  almost  always  contains 

CH3\ 

>CH  -  CH2  -  CH(NH2)  -  COOH   Leucine  (a-amino-isobutyl-acetic  acid), 
CH3/ 

.OH  (i)    Tyrosine  (p-oxyphenyl-a-aminopropionic 

C6H4<  •     acid) 

\CH2  -  CH(NH2)  -  COOH  (4) 
CH3  -  CH(OH)  -  COOH  Sarcolactic  acid  (dextro-lactic  acid). 

In  acute  phosphorus  poisoning  the  following  acids  can  be  detected  in  the  urine 
in  greatly  increased  quantity: 

/OH  (i) 

C6H4<\  Para-oxyphenyl-acetic  acid, 

\CH2  -  COOH  (4) 

,0H  (i)  Para-oxyphenyl-propionic  acid  (hydro- 

C6H4<(^  para-cumaric  acid). 

\CH2  -  CH2  -  COOH  (4) 
S  -  CH2  -  CHCNHs)  -  COOH 
Cystine,   |  ,  has  also  been  detected  in  phosphorus 

S-CH2-CH(NH2)-COOH     urine. 

In  phosphorus  poisoning  there  is  a  marked  decrease  in  the  urea-content  of 
the  urine  but  a  decided  increase  in  total  nitrogen.  A  considerable  part  of  the 
nitrogen,  that  is  to  say,  25  per  cent,  or  more  of  the  total  nitrogen,  appears  to  leave 
the  body  as  ammonia.  The  urine  of  adults  usually  contains  from  2  to  5  per  cent, 
of  the  total  nitrogen  as  ammonia.  The  increase  of  ammonia  may  have  some  con- 
nection with  the  increase  in  formation  of  acid  during  phosphorus  poisoning. 

Peptone-like  substances,  the  presence  of  which  is  attributed  to  profound  dis- 
turbance of  metabolism,  frequently  appear  in  the  urine  in  phosphorus 
poisoning.  Various  observers  believe  there  is  no  longer  any  doubt  as  to  the 
appearance  of  genuine  peptonuria.  A  glycosuria  may  also  appear,  or  be  easily 
induced  by  a  diet  rich  in  sugar.  In  accord  with  this  observation  is  the  fact  that 
the  liver  of  an  animal  poisoned  by  phosphorus  is  without  the  power  to  change 
glucose  of  the  blood  into  glycogen  and  store  up  the  latter.  In  phosphorus 
poisoning  the  alkalinity  of  the  blood  rapidly  diminishes  owing  to  the  increased 
formation  of  acid.  Since  persons  poisoned  bj'  phosphorus  have  icterus  (jaimdice), 
bile-pigment,  or  at  least  urobilin,  can  be  readily  detected  in  the  urine. 
2 


18 


DETECTION   OF   POISONS 


The  amounts  of  oxygen  and  carbon  dioxide,  which  the  organism  respectively 
takes  up  and  gives  off,  show  a  marked  diminution  during  phosphorus  poisoning. 
Only  48  per  cent,  of  carbon  dioxide,  as  compared  with  100  per  cent,  under  normal 
conditions,  may  be  eliminated. 

In  brief  the  chief  characteristics  of  phosphorus  urine  are  a  strong  acid  reaction; 
presence  of  protein  (peptone-like  substances);  and  frequently  occurrence  of  the 
amino  acids  mentioned  above,  as  well  as  fat  cylinders,  cell  detritus,  free  fat  glo- 
bules and  blood-corpuscles. 

Further  Examination  of  the  Distillate 

When  phosphorescence  has  been  distinctly  observed  in  the 
Mitscherlich  apparatus,  it  is  advisable  to  stop  distillation  and 
change  the  Liebig  condenser  to  its  customary  position.  This 
simpler  method  of  distilling  is  shown  in  Fig.  7  and  should  al- 
ways be  used  in  toxicological  analysis  when  there  is  no  occasion 
to  test  for  phosphorus. 


Fig.  7. — Distillation  with  Liebig  Condenser. 


Since  the  several  poisons  appearing  here  are  not  equally 
volatile  with  steam,  it  is  best  to  collect  the  distillate  in  two  or 
three  fractions.  The  first  will  contain  most  of  the  easily  volatile 
substances  Hke  hydrocyanic  acid,  chloroform,  ethyl  alcohol, 
acetone,  iodoform  and  nitrobenzene.  The  others  (second  and 
third)  will  contain  substances  less  easily  volatile  with  steam 
Hke  carbolic  acid,  aniline,  chloral  hydrate  and  carbon  disulphide. 
This  must  not  be  understood  to  mean  that  the  first  part  of  the 


VOLATILE   rOISONS  19 

distillate  will  be  free  from  substances  that  volatilize  with  diffi- 
culty, and  the  latter  part  free  from  those  that  volatilize  easily. 
In  the  main  such  will  be  the  separation,  but  either  part  of  the 
distillate  may  contain  traces  of  substances  which  will  appear 
in  larger  quantity  in  the  other  part. 

The  proper  procedure  is  to  distil  until  5  to  10  cc.  of  liquid 
have  been  collected.  Divide  the  distillate  into  several  portions 
and  test  for  hydrocyanic  acid,  chloroform,  ethyl  alcohol,  acetone, 
and,  if  necessary,  also  for  iodoform  and  nitrobenzene.  Use 
the  second  and  third  portions  (10  to  20  cc.)  to  test  for  carbolic 
acid,  aniline,  chloral  hydrate  and  carbon  disulphide. 

Several  of  these  volatile  substances  have  a  characteristic 
odor,  which  makes  it  possible  to  recognize  them  with  great 
certainty  in  the  original  material  and  especially  in  the  distil- 
late. First,  test  the  distillate  for  each  individual  substance 
by  its  most  characteristic  reaction.  Test  for  hydrocyanic 
acid  by  the  Prussian  blue  or  sulphocyanate  reaction;  for  ethyl 
alcohol,  a^cetone  and  acetaldehyde  by  Lieben's  reaction;  for 
carbolic  acid  and  aniline  by  Millon's  reaction;  for  chloroform, 
chloral  hydrate  and  iodoform  by  the  phenyliso cyanide  reaction; 
for  aniline  with  calcium  hypochlorite  solution;  and  finally,  for 
carbon  disulphide  with  lead  acetate  and  potassium  hydroxide 
solutions. 

When  there  is  reason  to  believe  that  a  certain  substance  is 
present,  confirm  the  result  by  making  other  characteristic  tests. 
It  is  seldom  necessary  to  examine  the  distillate  for  all  the  mem- 
bers of  the  group. 

HYDROCYANIC  ACID,  HCN 

Physiological  Action. — In  whatever  way  applied,  hydrocj^anic  acid  is  absorbed, 
even  from  the  skin.  So  rapid  is  the  absorption  of  this  poison  that  there  is 
evidence  of  an  intoxication  after  a  few  seconds,  or  a  few  minutes  at  most.  Part  of 
the  poison  thus  absorbed  passes  from  the  body  unchanged  by  way  of  the  lungs. 
Another  part,  usualty  much  less,  is  eliminated  by  the  kidnej's  and  passes  into  the 
urine.     Sweat  also  is  said  to  contain  Iwdrocj-anic  acid. 

Most  of  the  absorbed  hj'drocj-anic  acid,  though  variable  in  quantity,  undergoes 
chemical  change  within  the  organism  whatever  be  the  form  of  its  chemical  com- 
bination. H3-drocyanic  acid  is  supposed  to  combine  with  loosely  boimd  sulphur 
of  proteins  and  form  sulphocyanic  acid  (HSCN)  which  is  not  nearly  as  toxic  as 
hydrocyanic  acid.     (Antidote  for  hydrocyanic  acid.)     Hydrocyanic  acid  after  the 


20  DETECTION   OF   POISONS 

manner  of  the  cyanohydrin  reaction^  might  combine  chemically  with  carbohy- 
drates of  the  blood  and  tissues.  Finally,  putrefactive  changes  as  well  as  ferment 
action  within  the  cadaver  might  convert  hydrocyanic  acid  into  ammonium  for- 
mate.^ The  last  statement  may  explain  the  disappearance  of  hydrocyanic  acid 
until  only  traces  remain  in  the  cadaver.  Thus  the  possibility  of  making  more 
than  an  approximately  quantitative  determination  of  hydrocyanic  acid  taken 
internally  is  precluded  from  the  beginning.  Yet  there  are  instances  where  the 
poison  has  been  found  in  the  human  cadaver  after  14  days,  and  even  after  100  and 
180  days.  After  48  days  the  author  obtained  enough  hydrocyanic  acid  in  the  dis- 
tillate from  stomach  and  intestinal  contents  of  a  child  41/2  years  old  to  give  the 
Prussian  blue  test  in  three  different  portions  of  the  distillate  after  3  to  4  hours. 

Undoubtedly  hydrocyanic  acid  has  a  very  poisonous  effect  upon  ferments,  for 
it  kills  certain  vegetable  and  animal  enzymes,  or  at  least  strongly  retards  their 
action.  This  acid  interferes  particularly  with  the  action  of  that  enzyme  which 
causes  transfer  of  oxygen  from  blood-corpuscles  and  thereby  gives  rise  to  oxida- 
tive processes  (oxidation  ferment,  "respiration  ferment").  Careful  experiments 
in  metabolism  have  shown  that  warm-blooded  animals  under  the  influence  of 
hydrocyanic  acid  take  up  less  than  the  normal  amount  of  oxygen  and  con- 
sequently give  off  less  carbon  dioxide,  even  though  relatively  large  quantities  of 
oxygen  are  administered  artificially.  R.  Kobert  (Intoxikationen)  regards  hydro- 
cyanic acid  poisoning  as  an  internal  asphyxiation  of  the  organs  in  presence  of  an 
excess  of  oxygen.  The  oxidative  processes  of  the  blood  are  checked  and  so  little 
oxygen  is  used  that  the  venous  blood  becomes  arterial,  that  is  to  say,  contains  a 
large  quantity  of  oxyhsemoglobin.  As  a  result  the  color  of  the  venous  blood 
is  bright  red.  This  change  of  venous  to  arterial  blood  seems  to  be  permanent 
in  cold-blooded  but  usually  only  transitory  in  warm-blooded  animals.  The 
appearance  of  lactic  acid  in  the  blood  and  urine  is  due  to  the  disturbing  influence 
of  the  poison  upon  the  oxidative  processes  of  the  organism.  The  processes  of 
normal  metabolism  in  warm-blooded  animals  finally  oxidize  lactic  acid  to  car- 
bon dioxide  and  water.  Consequently  the  appearance  of  lactic  acid  in  the  blood 
is  very  transitory  and  it  is  not  found  in  the  urine  at  all.  The  occurrence  of  lactic 
acid  in  the  blood  and  a  decrease  in  its  alkalinity  are  concurrent.  As  a  result  of 
very  deficient  oxidation  during  hydrocyanic  acid  poisoning,  dextrose  not  infre- 
quently appears  in  the  urine. 

The  blood  therefore  in  hydrocyanic  acid  poisoning  is  characteristically  changed. 
Venous  blood  becomes  bright  red.  And  moreover  blood  which  contains  this  acid 
cannot  liberate  oxygen  from  hydrogen  peroxide,  that  is  to  say,  it  has  lost  its 
catalytic  power.'     Such  a  compound  as  cyano-haemoglobin  appears  to  exist  and 

1  R  -  c/      +  HCN  =  R  -  C(-OH  (R  denotes  any  radical) 

^O  \CN 

OH2  ^O 

2H-CN   =H-Cf 

O  H2  ^O  -  NH4 

^  Hydrocyanic  acid  poisons  platinum  black  just  as  it  does  blood  ferments.  Put 
about  5  cc.  of  3  per  cent,  hydrogen  peroxide  solution  in  each  of  two  test-tubes. 
Add  to  one  i  or  2  drops  of  hydrocyanic  acid  (about  i  per  cent,  solution)  and  to 
both  a  trace  of  platinum  black.  Pure  hydrogen  peroxide  at  once  gives  off  ox3^gen 
vigorously,  whereas  that  containing  hydrocyanic  acid  does  not. 


VOLATILE   POISONS  21 

its  formation  in  the  blood  of  a  person  poisoned  by  hydrocyanic  acid  would  seem 
probable,  yet  for  some  unknown  reason  the  union  of  this  acid  with  hsemoglobin 
takes  place  either  not  at  all  or  only  with  great  difficulty. 

In  a  chemical  examination  for  hydrocyanic  acid  and  potas- 
sium cyanide  the  contents  of  the  stomach  and  intestines,  or- 
gans rich  in  blood  as  liver,  brain  and  heart,  the  blood  itself 
and  sometimes  the  urine  are  most  important.  Examine  such 
material  at  once  for  hydrocyanic  acid  which  may  be  recognized 
by  its  characteristic  odor,  provided  putrefaction  has  not  gone 
too  far. 

Preliminary  Test. — A  special  test  (Schonbein-Pagenstecher 
reaction)  for  hydrocyanic  acid  should  precede  distillation. 
Acidify  a  portion  of  the  original  material  in  a  small  flask  with 
tartaric  acid  solution.  Then  suspend  in  the  flask  (see  Fig.  i)  a 
strip  of  "guaiac-copper"  paper^  without  letting  it  touch  the 
liquid.  Gently  warm  the  contents  of  the  flask  upon  the  water- 
bath.  Neither  hydrocyanic  acid  nor  potassium  cyanide  is 
present,  unless  the  paper  is  turned  blue  or  bluish  green.  But 
the  only  conclusion  to  be  drawn  from  a  positive  test  is  that 
hydrocyanic  acid,  or  an  easily  decomposable  cyanide,  may  be 
present.  Further  conclusions  should  not  be  drawn  from  a 
positive  result,  since  other  substances  like  ammonia,  volatile 
ammonium  compounds,  hydrochloric  acid  and  especially  oxi- 
dizing agents  Hke  ozone,  hydrogen  dioxide,  nitric  acid  and 
chlorine  will  turn  the  paper  blue.  Consequently  though  very 
delicate  this  test  cannot  be  accepted  as  conclusive  proof  of  the 
presence  of  hydrocyanic  acid. 

Mechanism  of  the  Reaction. — Hydrocyanic  acid  has  nothing  directly  to  do  with 
this  reaction.  But  it  forms  ozone  with  copper  sulphate  and  that  turns  the  guai- 
aconic  acid  of  guaiac  resin  blue.  Cupric  cyanide  (a)  is  an  intermediate  product 
which  furnishes  ozonized  oxygen  as  shown  in  (;8) : 

(a)  CUSO4        +  2HCN  =  Cu(CN)2  +  H2SO4, 

(/3)   6Cu(CN)2  +  3H2O    =  6CuCN    +  6HCN  +  O3. 

^Prepare  "guaiac-copper"  paper  by  saturating  strips  of  filter  paper  with 
freshly  prepared,  10  per  cent,  alcoholic  tincture  of  resin  of  guaiac.  Dr}-  these 
strips  in  air  and  moisten  before  using  with  very  dilute  aqueous  copper  sulphate 
solution  (i  :  1000). 


22  DETECTION   OF  POISONS 

The  actual  chemical  examination  for  hydrocyanic  acid  is  made 
by  adding  tartaric  acid  solution  to  the  finely  divided  material 
and  distilling  as  described  (see  page  i8).  This  acid  volatil- 
izes easily  with  steam  and  most  of  it  will  appear  in  the  first  part 
of  the  distillate.  Therefore  use  the  first  5  or  10  cc.  of  distillate 
for  the  tests.  Note  cautiously  the  odor  of  the  distillate,  which 
is  characteristic,  and  then  proceed  as  follows: 

1.  Prussian  Blue  Test. — Add  to  the  solution  (distillate) 
a  little  potassium  hydroxide  solution;  then  i  or  2  drops  of  fer- 
rous sulphate  solution  and  i  drop  of  ferric  chloride  solution. 
Shake  well  and  warm  gently.  Finally  acidify  with  dilute  hy- 
drochloric acid.  If  much  hydrocyanic  acid  is  present,  a  pre- 
cipitate of  Prussian  blue  will  appear  immediately.  But  if  the 
quantity  is  small,  the  solution  will  have  merely  a  blue  or  bluish 
green  color.  After  a  long  time  (10  to  12  hoars)  a  flocculent 
precipitate  of  Prussian  blue  will  settle  to  the  bottom  of  the  test- 
tube.     The  limit  of  delicacy  is  i  :  5,000,000.^ 

Mechanism  of  the  Reaction.  ^ — Hydrocyanic  acid  and  potassium  hydroxide 
form  potassium  cyanide  which  with  ferrous  sulphate  produces  ferrous  cyanide 
(a).  The  latter  combines  with  more  potassium  cyanide,  forming  potassium 
ferrocyanide  (0)  which  with  ferric  chloride  precipitates  Prussian  blue  (7),  the 
ferric  salt  of  hydroferrocyanic  acid  (H4Fe(CN)6). 

(a)  FeS04  +  2KCN  =  Fe(CN)2  +  K2SO4, 

(,8)   Fe(CN)2         +  4KCN  =  K4Fe(CN)6, 

(t)    3K4Fe(CN)6  +  4FeCl3  =  Fe4[Fe(CN)6]3  +  12KCI. 

Prussian  blue  will  not  appear  in  presence  of  alkalies,  since  they  decompose  it  as 
follows : 

Fe4[Fe(CN)6]3  +  12KOH  =  3K4Fe(CN)6  +  4Fe(OH)3. 
Consequently  test  the  final  mixture  with  blue  litmus  paper  to  make  sure  it  is  acid. 

2.  Sulphocyanate  Test. — Add  to  a  portion  of  the  distillate 
3  or  4  drops  of  potassium  hydroxide  solution  and  then  a  Httle 

1  Link  and  Meckel,' Zeitschrift  fiir  analytische  Chemie  17,  455  (1878). 

2  The  reaction  may  be  explained  by  saying  that  all  the  ferrous  salt  in  presence  of 
much  potassium  hydroxide  is  precipitated  as  ferrous  hydroxide  (a).  The  latter 
with  potassium  cyanide  then  forms  ferrous  cyanide  (/3)  and  finally  potassium 
ferrocyanide  (7): 

(a)  FeS04  +  2KOH  =  Fe(0H)2  +  K2SO4, 
(/3)  Fe(0H)2  +  2KCN  =  Fe(CN)2  +  2KOH, 
(7)    Fe(CN)2  +  4KCN  =  K4Fe(CN)6. 

With  ferric  chloride  potassium  ferrocyanide  in  presence  of  hydrochloric  acid 
finally  gives  Prussian  blue  (see  above). 


VOLATILE   POISONS  23 

yellow  ammonium  sulphide  solution.  Evaporate  to  dryness 
upon  the  water-bath.  Dissolve  the  residue  in  a  little  water, 
and  acidify  with  dilute  hydrochloric  acid.  Filter  through 
double  paper  to  remove  sulphur,  and  add  to  the  filtrate  2  or  3 
drops  of  ferric  chloride  solution.  If  the  distillate  contained 
hydrocyanic  acid,  a  reddish  to  blood-red  color  will  appear. 
This  is  due  to  ferric  sulphocyanate.  The  limit  of  delicacy  is 
I  :  4,000,000. 

Mechanism  of  the  Reaction. — Hydrocyanic  acid  and  potassium  hydroxide 
form  potassium  cyanide  which  takes  sulphur  from  yellow  ammonium  sulphide 
and  becomes  potassium  sulphocyanate  (a).  The  latter  with  ferric  chloride  forms 
ferric  sulphocyanate  (j8) : 

(«)  KCN      -f  (H4N)2Sx  =  KSCN        +  (H4N)2S._i, 
ip)   3KSCN  +  FeCls        =  Fe(SCN)3  +  3KCI. 

3.  Vortmann's^  Nitroprusside  Test. — Add  to  a  portion  of  the 
distillate  a  few  drops  of  potassium  nitrite  solution;  then  2  to  4 
drops  of  ferric  chloride  solution  and  enough  dilute  sulphuric 
acid  to  give  a  bright  yellow  color.  Heat  to  boiling,  add  suf- 
ficient ammonium  hydroxide  solution  to  remove  excess  of  iron 
and  filter.  Add  to  the  filtrate  i  or  2  drops  of  very  dilute  am- 
monium sulphide  solution.  If  the  solution  contained  hydro- 
cyanic acid,  a  violet  color  will  appear  and  pass  through  blue, 
green  and  yellow.     The  limit  of  delicacy  is  i  1312,000. 

Note. — ^This  test  is  the  reverse  of  the  nitroprusside  test  for  hydrogen  sulphide 
and  is  due  to  conversion  of  hydrocyanic  acid  to  potassium  nitroprusside,  K2Fe- 
(N0)(CN)6,  which  causes  the  color  changes  when  ammonium  sulphide  is  added. 
Very  small  quantities  of  hydrocyanic  acid  give  a  bluish  green  to  greenish  yellow 
color. 

4.  Silver  Nitrate  Test. — Acidify  a  portion  of  distillate  with 
dilute  nitric  acid,  and  add  silver  nitrate  solution  in  excess. 
If  hydrocyanic  acid  is  present,  a  white,  curdy  precipitate  of 
silver  cyanide  (AgCN)  will  appear.  Excess  of  ammonium 
hydroxide  solution  will  readily  dissolve  this  precipitate.  The 
limit  of  deKcacy  is  i  :  2.50,000. 

When  a  dilute  aqueous  solution  of  hydrochloric  acid  is  dis- 
tilled, the  acid  does  not  pass  into  the  distillate.  The  pre- 
cipitate,  therefore,   caused  by  silver  nitrate  solution  cannot 

^  Monatshefte  fiir  Chemie  7,  416  (1886). 


24  DETECTION   OF   POISONS 

possibly  be  silver  chloride,  because  a±  first  nothing  but  pure 
water  distils  from  a  i  per  cent,  or  weaker  solution  of  hydro- 
chloric acid.  To  rernove  hydrochloric  acid,  if  present,  re- 
distil the  first  distillate  over  borax.  This  will  retain  hydro- 
chloric but  not  hydrocyanic  acid.  It  is  advisable  to  collect 
upon  a  filter,  wash  and  dry  the  precipitate  caused  by  silver 
nitrate  solution.  If  silver  cyanide  is  heated  in  a  bulb-tube,  it 
will  form  metallic  silver  and  cyanogen  gas.  The  latter  may  be 
recognized  by  its  charactertisic  odor.  ^  The  reaction  is : 
aAgCN  =  2Ag  +  (CN)2. 

5.  Picric  Acid  Test. — Make  alkaline  a  portion  of  distil- 
late with  potassium  hydroxide  solution  and  heat  gently  (50  to 
60°)  with  a  few  drops  of  picric  acid  solution.  If  hydrocyanic 
acid  is  present,  the  solution  will  become  blood-red.  This  is  due 
to  formation  of  potassium  isopurpurate. 

Note. — This  test  is  not  as  delicate,  nor  as  characteristic  of  hydrocy  anic  acid 
as  the  other  tests.  If  hydrogen  sulphide  is  present,  as  it  frequently  is  in  distillates 
from  animal  matter,  picric  acid  solution  will  produce  a  red  color  owing  to  forma- 
tion of  picraminic  acid. 

6.  Weehuizen's^  Test. — Add  a  few  drops  of  phenolphthalin 
dissolved  in  dilute  sodium  hydroxide  solution  and  then  a  little 
copper  sulphate  solution  (i  :  2000)  to  a  portion  of  the  distillate. 
Even  in  a  dilution  of  i  :  500,000  hydrocyanic  acid  will  produce 
a  red  color  due  to  oxidation  of  phenolphthalin  to  phenolphthalein. 
Such  oxidizing  agents  as  hydrogen  peroxide,  nitric  acid  and 
ferric  chloride  do  not  give  this  test.  Paper  first  moistened  with 
alkahne  phenolphthalin  solution  and  then  with  very  dilute  copper 
sulphate  solution  may  be  used.  These  phenolphthaHn-copper 
sulphate  papers  turn  red  even  in  air  containing  hydrocyanic  acid. 

Under  the  conditions  of  the  test  phenolphthalin  is  oxidized  to  phenolphthalein: 


.--  "    H 
O  -f- 

'  ■- --  H 


/C6H4— OH  /C6H4— OH 

— C^C6H4— OH    =      C^C6H4— OH  +  H2O 

^C6H4  ^CbHi 


iO  — OC  O— CO 

Phenolphthalin  Phenolphthalein 

^  Owing  to  the  very  poisonous  character  of  cyanogen  gas,  it  is  safer  to  ignite  the 
gas  at  the  mouth  of  the  bulb-tube.  Cyanogen  gas  burns  with  a  purple  flame. 
Tr. 

2  Pharmaceutisch  Weekblad  42,  271;  and  Pharmaceutische  Zentralhalle  46, 
256  (1905). 


VOLATILE   POISONS  25 

On  the  other  hand  the  phlhalcin  heated  with  an  alkaline  hydroxide  and  zinc  dust 
is  reduced  to  the  phthalin. 

Quantitative  Estimation  of  Hydrocyanic  Acid 

To  determine  hydrocyanic  acid  quantitatively,  acidify  a  weighed  portion  of 
material  with  dilute  sulphuric  or  tartaric  acid  and  distil.  Determine  the 
quantity  of  hydrocyanic  acid  in  the  distillate  either  gravimetrically  or  volu- 
metrically.  If  the  former  method  is  used,  collect  the  precipitate  of  silver  cyanide 
upon  a  weighed  filter,  wash  and  dry  at  ioo°  to  constant  weight;  or  ignite  the 
precipitate  in  a  weighed  porcelain  crucible,  and  determine  the  quantity  of  me- 
tallic silver  obtained.  If  hydrochloric  acid  is  present  in  the  distillate,  redistil 
once  over  borax.     The  distillate  will  then  be  free  from  hydrochloric  acid. 

Detection  of  Hydrocyanic  Acid  in  Presence  of  Potassium  Ferrocyanide 

When  material  contains  non-poisonous  potassium  ferrocyanide,  hydrocyanic 
acid  will  appear  in  the  distillate  from  a  solution  acidified  with  tartaric  acid.  In 
an  experiment,  where  i  per  cent,  potassium  ferrocyanide  solution  was  distilled 
with  0.03  gram  of  tartaric  acid,  the  distillate  contained  considerable  hydrocyanic 
acid.  Carbon  dioxide,  passed  into  hot,  aqueous  potassium  ferrocyanide  solution, 
will  liberate  hydrocyanic  acid  even  at  water-bath  temperature  (75°).  To  test  for 
potassium  ferrocyanide  beforehand,  shake  some  of  the  original  material  with 
water  and  filter.  Test  the  filtrate  with  ferric  chloride  solution  and  dilute  hydro- 
chloric acid.  If  there  is  a  precipitate  of  Prussian  blue,  potassium  ferrocyanide  is 
present.  To  detect  free  hydrocyanic  acid,  potassium  or  sodium  cj'anide^  with 
certainty,  in  presence  of  potassium  ferrocyanide,  add  to  the  material  acid  sodium 
carbonate  in  not  too  small  quantity  and  distil.  Even  long  distillation  over  free 
flame  by  this  method  will  liberate  hydrocyanic  acid  only  from  simple  cj'anides  and 
not  from  potassium  ferrocyanide. 

Detection  of  Merciiric  Cyanide 

When  an  aqueous  solution  of  mercuric  cyanide,  which  is  exceedinglj'  poisonous, 
is  distilled  with  tartaric  acid,  the  distillate  will  contain  hydroc3'anic  acid  only 
when  a  large  quantity  of  mercuric  cyanide  is  present.  Distillation  of  100  cc.  of 
I  per  cent,  aqueous  mercuric  cyanide  solution  yields  a  distillate  which  gives  the 
Prussian  blue  test  distinctly.  But,  if  the  quantity  of  mercuric  cyanide  is  less  and 
the  solution  very  dilute  (for  example,  100  cc.  of  o.oi  per  cent,  solution),  there  will 
not  be  a  trace  of  hydrocyanic  acid  in  the  distillate,  even  though  the  solution  is 
strongly  acidified  with  tartaric  acid.  If,  however,  a  few  cc.  of  freshly  prepared 
hydrogen  sulphide  water  are  added  and  distillation  is  resumed,  mercuric  cj'anide 
will  be  completely  decomposed  and  the  distillate  will  contain  hydrocyanic  acid. 

Detection  of  Merciiric  Cyanide  in  Presence  of  Potassivun  Ferrocyanide 

The  method  of  detecting  hydrocyanic  acid  from  simple  cyanides,  in  presence  of 
potassium  ferrocj'anide,  is  not  applicable  to  mercuric  cyanide.  Long  distillation, 
even  from  saturated  acid  sodium  carbonate  solution,  gives  no  trace  of  hydrocj^aiuc 
acid.  But  distillation  in  presence  of  not  too  little  acid  sodium  carbonate,  after 
addition  of  a  few  cc.  of  freshly  prepared,  saturated  hj-drogen  sulphide  solution, 

^  Mercuric  cj'anide  is  an  exception. 


26  DETECTION   OF   POISONS 

liberates  hydrocyanic  acid  from  mercuric  cyanide  but  not  from  potassium 
ferrocyanide.  It  is  possible  to  detect  hydrocyanic  acid  by  this  method,  when 
very  little  mercuric  cyanide  is  mixed  with  considerable  potassium  ferrocyanide. 
For  example,  o.oi  gram  of  mercuric  cyanide  in  loo  cc.  of  lo  per  cent,  potassium 
ferrocyanide  solution  can  be  detected.  If  potassium  ferrocyanide  is  distilled 
directly  with  hydrogen  sulphide  without  addition  of  acid  sodium  carbonate, 
the  distillate  will  contain  considerable  hydrocyanic  acid. 

CARBOLIC  ACm 

Action  and  Fate  of  Carbolic  Acid  in  the  Animal  Body 

Concentrated  carbolic  acid  coagulates  and  destroys  the  constituents  of  the 

human  body,  especially  proteins  and  protoplasmic  structures.     It  has  therefore  a 

very  strong  caustic  action.     But  its  action  is  not  merely  local,  for  after  absorption 

OH  it  shows  an  affinity  particularly  for  the  central  nervous  system, 

1  brain  and  spinal  cord.     The  first  indications  of  this  in  animals  are 

/^\  strong  stimulation,  increased  irritability  as  in  the  case  of  strych- 

HC      CH       nine  and  paralysis.     In   man  the   period  of  stimulation  is   very 

I        11  slow  in  appearing.     In  chronic  poisoning,    after   repeated   small 

^  y  doses  of  carbolic  acid,  degeneration  of  the  kidneys  and  liver  is  a  re - 

Q  suit  of  absorption.     The  human  organism  absorbs  carbolic    acid 

H  very  rapidly.     Absorption   from    the  skin,  the    gastro-intestinal 

tract,  abrasions  and  the  respiratory  organs  takes  place  readily.     In  the  human 

organism  the  poison  is  converted  by  conjugation  with  sulphuric  acid  into  phenyl- 

sulphuric  acid: 

HO-SO2-OH  +  HO-CeHs  =  HO-SO2-OC6H5  +  H2O. 

When  the  quantity  of  carbolic  acid  is  very  large,  it  is  also  converted  into  phenyl- 
glycuronic  acid  by  conjugation  with  glycuronic  acid,  H00C-(CH.-0H)4CH0. 
Considerable  carbolic  acid  is  oxidized  within  the  body  to  dihydroxy-benzenes, 
namely  pyrocatechol  (C6H4(OH)2(i,2))  and  hydroquinol  (C6H4(OH)2(i,4)). 
These  enter  into  synthesis  with  sulphuric  acid  and  appear  in  urine  as  ethereal 
salts  of  sulphuric  acid.  The  dark  color  of  "carbolic  urine"  is  largely  due  to 
further  oxidation  of  hydroqmnol,  whereby  colored  products  (quinone  ?)  are 
formed.  In  carbolic  acid  poisoning,  urine  often  has  a  pronounced  dark  color 
(greenish  to  black).  Urine  in  other  cases  is  amber-yellow  at  first,  but  standing  in 
air  gives  it  a  deeper  color.  When  carbolic  acid  poisoning  is  suspected,  the  urine 
should  be  examined  chemically.  "Carbolic  urine"  differs  from  normal  human 
urine  in  being  nearly  free  from  sulphuric  acid,^  the  so-called  "preformed  sul- 
phuric acid."  Consequently  barium  chloride  solution,  in  presence  of  excess  of 
acetic  acid,  gives  only  a  slight  precipitate  of  barium  sulphate  or  none  at  all. 
Filter  when  there  is  a  precipitate  and  warm  the  clear  filtrate  with  a  few  cc.  of 
concentrated  hydrochloric  acid.  An  abundant  precipitate  of  barium  sulphate 
will  usually  appear.  The  mineral  acid  decomposes  phenyl-sulphuric  acid  into 
phenol  and  sulphuric  acid  which  is  then  precipitated.     Normal  human  urine 

^  This  is  sulphuric  acid  present  in  urine  as  sulphates.  It  is  also  termed  "  pre- 
formed sidphuric  acid,"  by  which  is  meant  that  it  enters  the  body  as  such.  In 
this  respect  it  differs  from  "ethereal,"  or  "conjugate"  sulphuric  acids,  which 
result  from  syntheses  within  the  body. 


VOLATILE   POISONS  27 

contains  considerably  more  "sulphate  sulphuric  acid"  (A — sulphuric  acid)  than 
"ethereal  sulphuric  acid"  (B — sulphuric  acid).  The  average  proportion  between 
the  two  being:  A — S04:B — S04=io:i.  Barium  chloride  solution,  added  to 
normal  urine  in  presence  of  acetic  acid,  produces  a  heavy  precipitate  of  barium 
sulphate. 

Distribution  of  Carbolic  Acid  in  the  Human  Body  After  Poisoning 

C.  Bischoff'  examined  organs,  removed  from  a  man  who  died  15  minutes  after 
taking  15  cc.  of  liquid  carbolic  acid,  and  found  the  poison  distributed  as  stated  in 
the  table  below.  The  organs  in  this  case  were  perfectly  fresh.  Only  a  small  por- 
tion of  the  stomach  was  received. 

Weight  Organ  Phenol 

242  grams  Contents  of  stomach  and  intestine  o. 171  gram 

112  grams  Blood  0.028  gram 

1480  grams  Liver  0.637  gram 

322  grams  Kidney  0.201  gram 

1445  grams  Brain  0.314  gram 

Bischoff  distilled  with  steam  until  the  distillate  gave  no  further  precipitate  with 
bromine  water.  The  results  show  how  rapidly  carbolic  acid  is  absorbed,  and  how 
soon  it  is  distributed  throughout  the  body. 

E.  Baumann*  has  published  certain  facts  relating  to  the  quantity  of  carboKc 
acid  formed  during  putrefaction  of  protein  substances.  Baumann  states  that  he 
obtained  from  100  grams  of  fresh  pancreas  and  100  grams  of  moist  fibrin,  mixed 
with  250  cc.  of  water,  after  6  days  of  putrefaction  0.073  to  0.078  gram  of  tri- 
bromophenol,  corresponding  to  0.0208  to  0.022  gram  of  phenol. 

Urine  gives  a  distinct  test  for  carbolic  acid  15  minutes  after  the  poison  has 
been  taken  by  the  mouth,  or  hypodermically.  This  shows  how  rapidly  carbolic 
acid  is  absorbed.  Most  of  the  carbolic  acid  absorbed  is  eliminated  in  4  or  5 
hours.  Schaffer^  found  the  quantity  of  conjugate  sulphuric  acid  in  urine  to 
increase  in  exact  proportion  to  the  quantity  of  carbolic  acid  taken. 

Tests  for  the  Detection  of  Carbolic  Acid 

Carbolic  acid  distils  quite  easily  with  steam.  Yet,  to  remove 
the  last  traces,  long-continued  distillation  is  necessary.  If 
fractional  distillation  is  made,  when  carboKc  acid  is  present, 
this  substance  will  appear  in  the  first  and  second  fractions  and 
even  in  the  third.  Usually  carboHc  acid  can  be  recognized  by 
its  peculiar  odor.  When  much  carbolic  acid  is  present,  the 
distillate  is  milky.     Colorless  or  reddish  globules  may  be  seen 

1  Berichte  der  Deutschen  chemischen  Gesellschaft  16,  1337  (1883). 

2  Berichte  der  Deutschen  chemischen  Gesellschaft  10,  685  (1877)  and  Zeitschrift 
fur  physiologische  Chemie  i,  61  (1877-78). 

'Journal  fur  praktische  Chemie,  Neue  Folge  18,  2S2  (1878). 


28  DETECTION   OF   POISONS 

floating  in  the  liquid.  Excess  of  potassium  or  sodium  hydroxide 
solution  will  dissolve  carbolic  acid  and  render  the  distillate 
perfectly  clear.  Pure,  anhydrous  carbolic  acid  melts  at  40 
to  42°  and  distils  at  178  to  182°. 

Decomposition  of  protein  substances  produces  phenol  and 
especially  para-cresoJ  in  small  quantity.  Traces  of  phenols 
can  almost  always  be  detected  in  distillates  from  animal  matter 
in  an  advanced  stage  of  decomposition.  Millon's  reagent, 
and  usually  bromine  water,  will  give  positive  tests  with  such 
distillates. 

I.  Millon's  Test. — Millon's  reagent,^  heated  with  a  solution 
containing  only  a  trace  of  carbolic  acid,  produces  a  red  color. 
An  aqueous  solution  containing  only  20  mg.  of  carbolic  acid, 
diluted  I  :  100,000,  will  give  a  distinct  red  color.  If  the  phenol 
solution  is  not  very  dilute,  the  color  will  appear  even  in  the  cold. 
Though  a  very  delicate  test,  it  is  not  characteristic  of  carbolic 
acid,  because  several  other  aromatic  compounds  behave  simi- 
larly.    This  is  true  of  derivatives  of  mon-acid  phenols  like  the 

three  cresols,  salicylic  acid,^  para- 
hydroxy-benzoic  acid,  para-hydroxy- 
phenyl-acetic  acid,  para-hydroxy- 
phenyl-propionic  acid  (hydro-para- 
cumaric  acid^)  and  tyrosine.  Aniline 
heated  with  Millon's  reagent  also 
gives  a  dark  red  color. 

2.  Bromine  Water  Test. — Excess 
of  bromine  water  produces  a  yellow- 
ish   white,     crystalline     precipitate, 
Fig.    8.-Tribromophenol     ^^^^  ^-^j^  ^-^^^^  CarboHc   acid 

Crystals.      From   a  dilution  .  . 

of  1 :  20  000.  solutions.     It  IS  a  very  delicate  test 

for  carbolic  acid.  Phenol  diluted 
I  :  50,000  yields,  after  some  time,  a  precipitate  made  up  in 
part  of  well-formed  crystals  (Fig.  8). 

1  For  the  preparation  of  this  reagent  see  page  314. 

^  Traces  of  salicylic  acid  volatilize  with  steam,  at  least  in  such  quantity  that  it 
can  be  detected  with  Millon's  reagent. 

^  Para-hydroxy-phenyl-acetic  acid  and  hydro-para-cumaric  acid  are  formed  in 
the  putrefaction  of  proteins  but  are  not  volatile  with  steam. 


VOLATILE    POISONS  29 

If  there  is  a  sufficient  excess  of  bromine  water  to  give  the  supernatant  liquid  a 
brownish  red  color,  the  precipitate  consists  only  of  tribromophenyl  hypobromite, 
C6H2Br40.  R.  Benedikt^  regards  this  compound  as  a  brom-phenoxy-tribromo- 
benzene  with  the  structure 

OBr         ,  whereas  Thiele  and  Eichwede^  have  ascribed  to  it  the  structure 
C  II 

/\  c 

BrC      CBr  /\ 

II       I  '                  BrC      CBr 

HC     CH  II       II 

X/  HC      CH 

C  \/ 

Br  C 

Br2 

This  reaction  takes  place  so  easily  that  carbolic  acid  may  even  be  determined 

quantitatively  as  this   tetrabromo-derivative  (see  page  31).     It  melts  at  132- 

OH  1^4°  with  evolution  of  bromine  and  crystallizes  as  lemon-yellow 

1  leaflets  from  alcohol-free  chloroform  or  ligroin.     Heated  with 

/'\  alcohol,  acetone,  xylene,  or  aqueous  sulphurous  acid,  this  com- 

BrCe^aCBr  pound  loses  bromine  and  changes  at  once  to  2,4,6-tribromo- 

I        II  phenol,   melting  at  93-94°.     Salicylic  alcohol  (saligenin),  sali- 

^  /  cylic  aldehyde,   salicylic  acid  and  para-hydroxy-benzoic    acid 

Q  are  converted  quantitatively  by  an  excess  of  saturated  bromine 

Br  water  even  in  the  cold  into  tribromo-phenyl  hypobromite. 


3.  Ferric  Chloride  Test. — Very  dilute  ferric  chloride  solution, 
added  drop  by  drop,  imparts  a  blue-violet  color  to  aqueous 
carbolic  acid  solutions.  Addition  of  dilute  hydrochloric  or 
sulphuric  acid  changes  this  color  to  yellow.  This  test  is  not  as 
delicate  as  i  and  2.  It  is  entirely  negative  in  presence  of  min- 
eral acids.     The  limit  of  dehcacy  is  about  i  :  1000. 

4.  Hjrpochlorite  Test. — Add  a  few  cc.  of  ammonium  hydrox- 
ide solution  to  a  dilute,  aqueous  carbolic  acid  solution,  and  then 
2  or  3  drops  of  freshly  prepared  calcium  or  sodium  hypochlorite 
solution.  Gentle  warming  will  produce  a  blue  color.  Very 
dilute  carbolic  acid  solutions  after  some  time  give  only  a  green 
to  blue-green  color.  F.  A.  Fluckiger^  allows  bromine  vapor 
to  come  into  contact  with  the  phenol  solution  which  has  been 
mixed  with  a  little  ammonium  hydroxide  solution  in  a  porcelain 
dish. 

^  Annalen  der  Chemie  und  Pharmazie  199,  127  (1879). 
^Berichte  der  Deutschen  chemischen  Gesellchaft  33,637  (1900). 
^  Pharmaceutische  Chemie,  page  2S7  (1S79). 


30  DETECTION   OE   POISONS 

5.  Nitrite  Test. — Mix  a  carbolic  acid  solution  with  a  dilute 
alcoholic  solution  of  ethyl  nitrite,  C2H5-0-N  =  0/  or  iso- 
amylnitrite,  C5Hii-0-N  =  0,^  and  add  concentrated  sulphuric 
acid  from  a  pipette  so  that  it  forms  a  distinct  under-layer.  A 
red  zone  will  appear  at  the  contact  surface  of  the  two  liquids. 
This  is  a  very  delicate  test. 

This  test  may  also  be  made  by  adding  the  Hquid  under  ex- 
amination as  an  upper  layer  upon  concentrated  sulphuric  acid 
containing  a  trace  of  red  fuming  nitric  acid. 

6.  H.  Melzer's  Benzaldehyde  Test.^ — Add  2  cc.  of  concen- 
trated sulphuric  acid  to  i  cc.  of  the  solution  (distillate)  to  be 
tested  for  carboHc  acid,  then  i  or  2  drops  of  benzaldehyde  and 
heat.  The  mixture,  at  first  yellowish  brown,  will  become  dark 
red.  At  the  same  time  a  red  resinous  substance  will  appear, 
unless  the  solution  is  too  dilute.  When  cold  add  10  cc.  of  water 
and  enough  potassium  hydroxide  solution  to  give  a  distinct 
alkaline  reaction.  If  carbolic  acid  is  present,  a  violet-blue 
color  will  appear.  To  obtain  this  coloring-matter,  acidify  the 
solution,  extract  with  ether  and  evaporate  the  solvent.  Alka- 
lies, added  to  alcoholic  solutions  of  the  coloring-matter,  produce 
a  blue  color  which  acids  discharge.  This  is  a  very  dehcate 
test.  One  cubic  centimeter  of  0.05  per  cent,  carbolic  acid  solu- 
tion (=  0.0005  gram  of  carbolic  acid),  will  still  give  the  blue 
color  very  distinctly. 

Note. — In  absence  of  phenol  concentrated  sulphuric  acid  produces  a  dark 
brown  color  with  benzaldehyde.  According  to  A.  Russanow*  the  first  condensa- 
tion product  between  phenol  and  benzaldehyde  in  presence  of  concentrated 
sulphuric  acid  is  para-dihydroxy-triphenyl-methane  which  crystallizes  in  yellow- 
ish needles: 

CeHfiv  ---"  'HiC6H4-OH  C6H5  C6H4-OH 

\c=iO+     i  =H20+  >C<  (1,4). 

h/  .-,..    HIC6H4-OH  H-^     \eH4-OH 

Benzaldehyde  Phenol  p-Dihydroxy-triphenyl-methane 

Akalies  dissolve  the  pure  crystals  without  color  but,  if  these  solutions  are 
exposed  to  air,  oxidation  takes  place  and  a  red  or  red- violet  color  appears.     Prob- 

^  The  officinal  preparation  is  called  "  Spiritus  Aetheris  Nitrosi." 

2  Amylium  nitrosum  of  pharmacists. 

2  Zeitschrift  fur  analytische  Chemie  37,  345  (1898). 

^  Berichte  der  Deutschen  chemischen  Gesellschaft  22,1943  (i 


VOLATILE   POISONS  31 

ably  benzaurine,  dihydroxy-triphcnyl-carbiiiol,  is  first  formed.     This  compound 
is  a  brick-red  crystalline  powder  soluble  in  alkalies  with  a  violet  color. 

QUANTITATIVE  METHODS  OF  ESTIMATING  PHENOL 

1.  Gravimetric  Estimation  as  Tribromophenol 

The  principle  of  this  method  is  based  on  the  complete  pre- 
cipitation of  phenol  from  aqueous  solution  as  tribromophenyl 
hypobromite  by  an  excess  of  saturated  bromine  water  (see 
Test  2) .  The  precipitate  is  practically  insoluble  in  cold  bromine 
water  and  the  results  are  very  satisfactory.^ 

Procedure. — Place  the  aqueous  phenol  solution  in  a  large 
glass-stoppered  flask.  Add  gradually,  while  shaking,  saturated 
bromine  water  until  the  supernatant  liquid  has  a  red-brown 
color  and  bromine  vapor  is  visible  above  the  solution.  Let 
stand  2-4  hours  and  shake  frequently.  Then  collect  the  pre- 
cipitate in  a  weighed  Gooch  crucible  and  dry  in  a  vacuum  des- 
iccator over  sulphuric  acid  to  constant  weight.  On  the  basis  of 
the  following  proportion  calculate  the  weight  of  phenol  corre- 
sponding to  the  weight  of  the  precipitate: 

C6H2Br40  :  C^Hc-OH  =  Wt.  of  Ppt.  found  :  x 
409.86  94.05 

Since  the  ratio  — —z^  =  0.2295,  the  weight  of  phenol  may  be 
found  by  multiplying  the  weight  of  the  precipitate  by  0.2295. 

2.  Beckurts-Koppeschaar^  Volvmietric  Method 

Dilute  sulphuric  acid  liberates  hydrobromic  acid  from  potas* 
slum  bromide  (a)  and  bromic  acid  from  potassium  bromate  {^). 
These  two  acids  react  according  to  (7) : 

(a)  KBr      +  H.SO4   =  KHSO4  4-  HBr, 
(;8)   KBrOs  +  H2SO4  =  KHSO4  +  HBrOs, 
(7)    sHBr    +  HBrOs  =  sBra       +  3H2O. 

1  The  following  results  were  obtained  by  F.  Beuttel: 

Phenol  taken  C6H2Br40  Phenol  found  Per  cent,  found 

1.  0.103    grm.  0-4538  grm.  0.0997  grm.  96.2 

2.  0.2072  grm.  0.8S06  grm.  0.2014  grm.  98.6 

3.  0.2072  grm.  0.8708  grm.  0.2006  grm.  98.6 

2  Archiv  der  Pharmazie  24,  570  (1886). 


32  DETECTION   OP   POISONS 

Therefore  addition  of  dilute  sulphuric  add  to  a  mixture  of 
potassium,  bromide  and  bromate  solutions  liberates  bromine 
which  will  convert  phenol  into  a  mixture  of  tribromophenol  and 
tribromophenyl  hypobromite.  The  excess  of  free  bromine 
and  also  the  loosely  bound  bromine  atom  of  tribromophenyl 
hypobromite  will  displace  iodine  from  potassium  iodide  and 
finally  all  the  phenol  will  be  present  as  tribromophenol: 

CeHzBrsOBr  +  2KI  =  CeHaBrgOK  +  KBr  +I2 

One  molecule  of  phenol  requires  6  atoms  of  bromine,  as  shown 
by  the  equation: 

SKBr  +  KBrOs  +  6H2SO4  +  CcHbOH  =  CeHsBrsOH  +  sHBr  +  6KHSO4  + 

3H2O. 

The  following  standard  solutions  are  required: 

I.  o.oi    n-potassium    bromide    solution,    containing 


100 


grams  =  '—  =  5.956  grams  KBr  in  1000  cc. 

iKBrOs 

2.  0.01   n-potassium  bromate  solution,  containing  

167.17 
grams  =  =  1.6717  grams  KBrOs  m  1000  cc. 

3.  0.1  n-sodium  thiosulphate  solution,  containing  o.i 
Na2S203.5H20  grams  =  24,83  grams  in  1000  cc. 

4.  Potassium  iodide  solution,  containing  125  grams  of  KI 
in  1000  cc. 

Procedure. — Put  about  25  cc.  of  aqueous  phenol  solution 
(distillate)  into  a  flask  having  a  tight  glass  stopper.  Add  50 
cc.  each,  of  o.oi  n-potassium  bromide  and  o.oi  n-potassium  bro- 
mate solutions,  then  5  cc.  of  pure  concentrated  sulphuric  acid 
and  shake  vigorously  for  several  minutes.  The  gradually 
increasing  opalescence  of  the  solution  becomes  more  and  more 
marked,  as  tribromophenol  and  tribromophenyl  hypobromite 
are  precipitated.  The  yellow  color  which  soon  appears  shows 
excess  of  bromine.  Open  the  flask  in  15  minutes,  add  10  cc. 
of  potassium  iodide  solution,  shake  and  titrate  free  iodine  in 
5  minutes  with  o.i  n-sodium  thiosulphate  solution. 


VOLATILE    POISONS  33 

6  gram-atoms  Br       6  X  79.96 

Calculation. — = =  4.7976  grams  of  bromine  are 

100  100 

set  free  from  a  mixture  of  1000  cc.  of  o.oi  n-potassium  bromide  solution  and  1000 
cc.  of  0.01  n-potassium  bromatc  solution.     A  mixture  therefore  of  50  cc.  of  each 
of  the  two  solutions  will  give  0.2399  gram  of  bromine.     This  quantity  of  bro- 
mine can  convert  0.04704  gram  of  phenol  into  tribromophenol: 
6Br :  C0H5OH 
479.76     94.05     =  0.2399  :  X  (x  =  0.04704) 

I  cc.  of  0.1  n-sodium  thiosulphate  solution  corresponds  to  0.012697  gram  of 
iodine  and  this  quantity  of  iodine  to  0.007996  gram  of  bromine.  But  0.007996 
gram  of  bromine  will  convert  0.00157  gram  of  phenol  into  tribromophenol: 

6Br :  CoHeOH 

479.76     94.05      =  0.007996  :  X  (x  =  0.00157) 

Consequently,  for  each  cc.  of  o.i  n-sodium  thiosulphate  solution  used,  subtract 
0.00157  from  0.04704  gram  of  phenol.  This  determines  the  quantity  of  car- 
bolic acid  in  the  25  cc.  of  distillate  taken. 

3.  Messinger-Vortmann^  Volumetric  Method 
Excess  of  iodine  (8  atoms  of  iodine  to  i  molecule  of  pheno 
dissolved  in  4  molecules  of  potassium  hydroxide),  added  to  an 
alkaline  phenol  solution  at  50-60°,  will  produce  a  dark  red,  non- 
crystalline precipitate.  One  molecule  of  phenol  requires  6 
atoms  of  iodine: 

1.  CeHsOH  +      3I2     =  C6H2I3OH  -f  3HI, 

2.  3HI  +  3KOH  =       3H2O       +  3KI. 

This  red  precipitate  dissolves  in  hot  potassium  hydroxide 

solution  with  a  red-brown  color  and  appears  as  white  2,  4,  6-tri- 

iodophenol,  melting  at  154-156°,  on  addition  of  an  excess  of 

dilute  sulphuric  acid.     Messinger  and  Vortmann  regard  the 

red  compound  as  di-iodophenyl  hypoiodite  (C6H3I2OI)  which 

potassium  hydroxide  converts  into  the  more  stable  isomeric  tri- 

iodophenol: 

01  OH 


c 

C 

/\ 

is  converted    1 

by       /\ 

IC      CI 

potassium  hy- 

IC     CI 

1     II 

droxide 

into 

1          1 

HC      CH 

HC      CH 

\/ 

\/ 

c 

C 

H 

I 

Red  di-iodo- 

White 2,  4,  6-tri. 

phenyl  hypo-iodite 

iodophenol 

^  Berichte  der  Deutschen  chemischen  Gesellschaft  22,  2312  (1S89);  and  23, 
2753  (1890).     See  also  Kossler  and  Pennj^  Zeitschrift  fur  physiologische  Chemie 
17,  117  (1892). 
3 


34  DETECTION   OE  POISONS 

Prpcediire.' — The  reaction  between  the  alkaline  phenol  solu- 
tion and  iodine  is  rather  slow  in  the  cold  but  is  hastened  at  50 
to  60°. 

Place  a  measured  volume  of  aqueous  phenol  solution  (5  to 
10  cc.)  in  a  small  flask  and  add  a  measured  volume  of  o.i 
n-potassium  hydroxide  solution  until  the  mixture  is  strongly 
alkaline.  Warm  gently  by  dipping  the  flask  in  water  at  60° 
and  add  10-15  cc.  more  of  o.i  n-iodine  solution  than  the 
volume  of  0.1  n-potassium  hydroxide  solution  used,  or  until  the 
excess  of  iodine  produces  a  strong  yellow  color.  Agitation  will 
cause  a  deep  red  precipitate  to  appear.  Cool  the  solution, 
acidify  with  dilate  sulphuric  acid  and  dilute  to  a  definite  volume 
(250  to  500  cc.) .  Filter  an  ahquot  portion  (100  cc.)  rapidly  and 
determine  excess  of  iodine  with  0.1  n-sodium  thiosulphate  solu- 
tion. 

Calculation. — Each  molecule  of  phenol  requires  6  atoms  of 

rru      ,               .           f    ■  A'           CeHsOH       94.05 
iodine.     Therefore    i    atom   of    lodme  = 7 =  — 7 —  = 

15.675  phenol.  1000  cc.  of  0.1  n-iodine  solution,  containing 
O.I  gram-atom  of  iodine,  correspond  therefore  to  1.5675  grams 
of  phenol. 

Note. — This  method  will  not  give  satisfactory  results,  unless 
at  least  3  molecules  of  sodium  or  potassium  hydroxide  are  taken 
for  I  molecule  of  phenol. 

Estimation  of  Phenol  in  Urine 

In  determining  carbolic  acid  in  urine,  the  regular  occurrence  of  phenols  must 
not  be  overlooked.  After  a  mixed  diet,  the  quantity  of  normal  human  urine 
passed  in  24  hours  will  yield  approximately  0.03  gram  of  phenols  (phenol  and 
more  especially  para-cresol). 

In  certain  diseases  where  there  is  excessive  bacterial  decomposition  within  the 
organism,  in  the  intestines  for  example,  urine  contains  more  of  these  phenols  and, 
consequently,  more  conjugate  sulphuric  acids.  Even  external  application  of 
carbolic  acid,  for  instance  the  use  of  carbolic  acid  water  as  a  lotion,  is  sufficient  to 
increase  the  quantity  of  phenyl-sulphuric  acid  in  urine. 

Detection  of  Carbolic  Acid  in  Presence  of  Aniline 

Aniline  closely  resembles  carbolic  acid  in  behavior  toward  Millon's  reagent  and 
bromine  water.     But  the  two  substances  can  be  easily  separated.     Add  potassium 

^  Use  0.5  to  I  per  cent,  carbolic  acid  solution  for  laboratory  experiments. 


VOLATILE   POISONS  35 

hydroxide  solution  in  large  excess  and  distil.  The  distillate  will  contain  aniline 
alone.  Or  make  the  solution  strongly  acid  with  dilute  sulphuric  add, 
and  extract  with  ether  which  will  dissolve  only  carbolic  acid.  Evaporate 
the  ether  extract  at  a  moderate  temperature  and  examine  the  residue. 

CHLOROFORM 

Behavior  in  the  Human  Organism. — When  inhaled  chloroform  first  passes  from 
the  air  into  the  blood-plasma  which  then  transmits  it  to  the  red  blood-corpuscles 
TT  where  it  may  accumulate  in  relatively  large  quantity.     Air  passed 

I  through   blood   will   remove   chloroform   completely.     Pohl  (see 

CI — C — CI  Robert's  "Intoxikationen")  states  that  blood  may  contain  0.62 
I  per  cent,  of  chloroform,  three-fourths  of  which  will  be  in  the  red 

blood-corpuscles.  At  the  height  of  a  harmless  narcosis  the  blood 
contained  only  0.035  per  cent,  of  chloroform.  Absorption  of  chloroform  is  rapid 
from  all  parts  of  the  body.  The  stimulative  action  of  chloroform  on  the  mucous 
membranes  of  the  respiratory  passages  explains  such  disturbances  as  coughing, 
secretion  of  saliva  and  reflex  slowing  of  respiration  and  heart-beat,  occurring  at 
the  beginning  of  narcosis.  Dilatation  of  the  blood-vessels  of  organs  living  after 
death  is  due  to  paralysis  caused  by  even  small  doses  of  chloroform.  A  drop  in 
blood-pressure  accompanies  paralysis  of  the  brain  and  the  heart's  action  is  feebler 
and  slower.  Several  researches  regarding  the  effect  of  inhaled  chloroform  upon 
human  and  animal  metabolism  have  shown  an  increase  in  the  quantity  of  nitrogen 
in  the  urine  after  prolonged  narcosis  because  more  protein  is  decomposed.  The 
amount  of  neutral  sulphur  and  chlorine  in  the  urine  also  increases.  The  increase 
of  the  latter  is  due  in  part  at  least  to  the  conversion  of  chloroform  into  chloride. 
The  acidity  of  the  urine  is  also  much  higher.  A  final  characteristic  of  chloroform 
urine  is  the  high  content  of  reducing  substances.  The  increased  protein  decom- 
position in  chloroform  narcosis  affects  both  reserve  protein  and  that  of  the  tissues. 
This  may  explain  degeneration  in  red  blood-corpuscles,  glandular  organs,  the 
heart,  etc.,  after  frequent  narcoses  or  one  of  long  duration. 

The  temporary  or  permanent  paralysis  of  isolated  animal  or  vegetable  cells, 
such  as  leucocytes,  ciliated  cells,  yeast  cells,  algae  and  spores,  is  evidence  of  the 
antiseptic  action  of  chloroform  when  present  in  proper  concentration  in  air  or  in  a 
liquid.  This  explains  the  use  of  i  per  cent,  aqueous  chloroform  solution  as  an 
antiseptic.  Added  to  urine  it  acts  as  a  preservative.  Therefore  it  may  be  used 
in  the  study  of  the  action  of  enzymes  but  not  of  bacteria,  though  all  micro-organ- 
isms are  not  paralyzed  or  killed  by  chloroform  water. 

Pohl  and  Hans  Meyer  have  studied  the  distribution  of  chloroform  in  the  body 
and  found  that  the  red  blood-corpuscles  and  the  brain  are  most  likely  to  show  this 
poison.  After  chloroform  has  been  inhaled,  some  will  appear  in  the  gastric  Juice 
but  at  most  only  traces  in  the  urine.  In  but  two  out  of  15  cases  of  chloroform 
narcosis  was  this  poison  found  in  the  urine  and  then  onl}^  in  traces. 

Kobert  states  that  as  a  rule  it  is  the  exception  to  find  chloroform  itself  in  the 
cadaver,  because  part  of  the  poison  is  converted  into  chloride  in  the  human  organ- 
ism and  part  is  quickly  exhaled  during  respiration.  Usually  it  is  possible  to  detect 
chloroform  in  the  breath  of  patients  even  24  hours  after  narcosis.  Budinger 
states  that  the  mucus  of  the  respiratory  passages  retains  chloroform. 


36  DETECTION   OE   POISONS 

Tests  for  the  Detection  of  Chloroform 

Chloroform  distils  easily  with  steam  and  appears  in  the  first 
fraction  in  largest  quantity.  When  much  chloroform  is  present, 
it  will  separate  from  the  distillate  as  heavy,  colorless  globules, 
whereas  a  small  quantity  will  remain  in  solution.  This  solution 
usually  has  the  characteristic  odor  and  sweetish  taste  of  chloro- 
form. The  following  tests  should  be  applied  to  the  first  frac- 
tion. 

1.  Phenylisocyanide  Test. — Add  i  or  2  drops  of  aniline  to  the 
chloroform  solution  (distillate),  and  then  a  few  cc.  of  aqueous, 
or  alcohoHc  potassium  hydroxide  solution.  Gentle  heat  will 
produce  phenyUsocyanide  (CeHgNC).  The  penetrating  and 
very  repulsive  odor  of  this  compound  is  easily  recognized. 

CHCI3  +  CeHs-NHa  +  3KOH  =  CeHg.NC  +  3KCI  +  3H2O. 

A.  W.  Hofmann  states  that  this  test  will  show  with  certainty 
I  part  of  chloroform  in  5000  to  6000  parts  of  alcohol. 

Note. — Chloral,  chloral  hydrate,  bromoform,  iodoform  and  tetrachloro- 
methane  also  give  this  test. 

The  fact  that  aniline  boiled  with  potassium  hydroxide  solution  gives  a  peculiar, 
faintly  ammoniacal  odor,  even  when  chloroform  is  absent,  must  not  be  over- 
looked. There  is  small  chance,  however,  of  confusing  this  odor  with  the  repulsive 
smell  of  phenylisocyanide.  In  doubtful  cases  warm  some  water,  containing  a 
drop  of  aniline  and  a  trace  of  chloroform,  with  potassium  hydroxide  solution  and 
compare  the  odor  with  that  in  question. 

2.  Schwarz's  Resorcinol  Test.^ — Dissolve  about  o.i  gram  of 
resorcinol  ( C6H4  \'  ^jr  )  n  )  in  2  cc.  of  water,  add  a  few  drops  of 

sodium  hydroxide  solution  and  finally  the  hquid  containing 
chloroform.  This  mixture  heated  to  boiling  will  develop  even 
in  very  dilute  solution  a  yellowish  red  color  attended  by  a 
beautiful  yellowish  green  fluorescence. 

Chloral,  bromal  bromoform  and  idoform  also  give  this  test. 

3.  Lustgarten's-  Naphthol  Test. — Dissolve  a  few  centigrams 
of  a-  or  jS-naphthol  in  i  or  2  cc.  of  33  per  cent,  aqueous  potas- 
sium hydroxide  solution.     Warm  to  50°  and  add  the  solution  to 

^  Zeitschrift  fiir  analytische  Chemie,  27,  668. 
^Monatshefte  fiir  Chemie,  3,  715  (1882). 


VOLATILE   POISONS  37 

be  tested.  Chloroform  will  produce  an  evanescent  blue  color 
which  in  contact  with  air  will  change  to  green  and  then  to 
brown.  This  color  is  less  stable  when  /3-naphthol  is  used. 
Acidification  of  the  blue  solution  will  precipitate  naphthol  col- 
ored by  a  red  dye  stuff.  This  precipitate  is  usually  brick-red. 
Chloral,  bromal  bromoform  and  idoform  also  give  this  test. 

4.  Cyanide  Test. — Seal  the  Hquid  to  be  tested  for  chloroform 
in  a  glass  tube  (pressure-tube^)  with  a  little  soHd  ammonium 
chloride  and  alcohohc  potassium  hydroxide  solution.  Heat 
for  several  hours  in  a  boiling  water-bath.  Cool  the  tube,  re- 
move the  solution  and  test  for  hydrocyanic  acid  by  the  Prussian 
blue  reaction.  A  positive  test  means  that  the  distillate  con- 
tained chloroform.     The  following  reactions  take  place: 

(a)  CHCI3  +  H3N  +  3KOH  =  HCN  +  3KCI  +  3H2O, 
(^)  HCN  +    KOH  =  KCN  +  H2O. 

5.  Reduction  Tests,  (a)  "With  Fehling's  Solution. — Warm 
the  liquid  containing  chloroform  with  Fehling's  solution.  A 
red  precipitate  of  cuprous  oxide  will  appear. 

(b)  With  Ammoniacal  Silver  Nitrate  Solution. — Add  excess 
of  ammonium  hydroxide  to  silver  nitrate  solution  and  then 
the  liquid  containing  chloroform.  Heat  will  produce  a  black 
precipitate  of  metallic  silver. 

These  reactions  are  not  characteristic  of  chloroform,  because 
many  volatile  organic  subtances,  as  formic  acid  and  aldehydes 
which  may  occur  in  distillates  from  animal  material,  reduce 
Fehling's  and  ammoniacal  silver  nitrate  solutions. 

Quantitative  Estimation  of  Chloroform  in  Cadavers 

(Ludwig-Fischer^) 

Mix  a  weighed  portion  of  material  with  water  and  distil 
as  long  as  there  is  any  chloroform.  To  tell  when  this  point  is 
reached,  apply  the  phenylisocyanide  test  to  a  few  cc.  of  liquid 

^  An  ordinary  citrate  of  magnesium  bottle  is  a  convenient  apparatus  for  this 
test.  Wrap  a  towel  around  the  bottle,  place  it  in  the  water-bath  and  gradually 
raise  the  temperature  to  boiling.     Do  not  remove  the  bottle  until  it  is  cold.     Tr. 

2  Jahresbericht  des  chemischen  Untersuchungsamtes  der  Stadt  Breslau  fiir  die 
Zeit  vom  i  April  1804  bis  31  ]\Iarz  1S05. 


38  DETECTION   OF   POISONS 

collected  at  the  end  of  distillation.  Add  some  calcium  car- 
bonate to  combine  with  free  hydrochloric  acid.  Warm  the 
distillate  to  about  60°  and  draw  washed  air  through  it  by  suc- 
tion. Pass  this  air  through  a  combustion-tube  heated  to  high 
temperature  and  then  into  silver  nitrate  solution  acidified  with 
nitric  acid.     Weigh  the  precipitated  AgCl  (N). 

Calciilation : 

sAgCl :  CHCI3  =  N  :  X. 

This  method  is  based  upon  the  fact  that  chloroform  heated 

with  steam  above  200°  is  decomposed  into  carbon  monoxide, 

hydrochloric  and  formic  acids : 

(a)  CHCI3  +  H2O    =  CO  +  3HCI. 

(/3)   CHCI3  +  2H2O  =  H.COOH  +  3HCI. 

In  a  series  of  blank  experiments  B.  Fischer  has  shown  that  the  stomach, 
stomach  contents  and  blood,  of  a  person  who  has  not  taken  chloroform,  give 
no  volatile  chlorine  compounds  under  these  conditions.  By  this  method  B. 
Fischer  found  in  the  cadaver  of  a  laborer,  who  had  died  during,  chloroform 
narcosis,  the  following  quantities  of  chloroform: 


Weight 

Organ 

Chloroform 

985  grams 

Stomach  and  contents  and  parts  of 

the  intestine 

0.1      gram 

780  grams 

Lungs  and  blood  from  the  heart 

O.OS5  gram 

445  grams 

Portions  of  spleen,  kidneys  and  liver 

traces 

480  grams 

Brain 

0.07     gram 

From  these  results  it  appears  that  most  of  the  chloroform  was  in  the  brain  and 
blood. 

CHLORAL  HYDRATE 

CI 

I  Chloral  hydrate  distils  very  slowly  v/ith  steam  from  an  acid 

I  solution.    Therefore  the  complete  distillation  of  a  large  quantity 

jj (2 OH     °^  chloral  hydrate  requires  considerable  time.     Chloral  hydrate 

appears  as  such  in  the  distillate. 


OH 


Tests  for  the  Detection  of  Chloral  Hydrate 


Chloral  hydrate  like  chloroform  will  give  the  phenyliso- 
cyanide,  resorcinol  and  Lustgarten's  naphthol  tests.  But  the 
distillate  containing  chloral  hydrate  does  not  have  the  charac- 
teristic chloroform  odor  which  is  also  scarcely  perceptible  in 
very  dilute  aqueous  chloroform  solutions. 


VOLATILE   POISONS  39 

Jaworowski^  suggests  the  following  tests  to  differentiate 
chloral   hydrate   from    chloroform: 

1.  Test  with  Nessler's  Solution. — Add  a  few  drops  of  this  re- 
agent to  an  aqueous  chloral  hydrate  solution  and  shake.  It 
will  produce  a  yellowish  red  precipitate,  the  color  of  which  will 
change  after  a  while  to  a  dirty  yellowish  green.  This  is  an 
aldehyde  reaction. 

2.  Test  with  Sodium  Thiosulphate. — Boil  a  few  cc.  of  chloral 
hydrate  solution  with  0.2-0.3  gram  of  solid  sodium  thiosulphate. 
This  will  give  a  turbid  liquid  of  brick-red  color.  A  few  drops 
of  potassium  hydroxide  solution  will  remove  the  turbidity  and 
change  the  color  to  brownish  red. 

When  the  quantity  of  chloral  hydrate  is  not  too  small,  it  may 
also  be  detected  by  the  following  procedure: 

Decomposition  of  Chloral  Hydrate. — Heat  a  portion  of  the 
distillate  for  30  minutes  under  a  reflux  condenser  with  calcined 
magnesium  oxide  (MgO)  upon  a  boiling  water  bath.  Magne- 
sium formate  and  chloroform  are  produced  by  decomposition 
of  chloral  hydrate. 

2CCl3.CH(OH)2  +  MgO  =  2CHCI3  +  Mg(00CH)2  4-  H2O. 

Proceed  as  follows  to  detect  these  products: 

Chlorofomi. — Distil  a  few  cc.  from  the  solution  in  the  flask 
and  test  for  chloroform  by  the  phenyKsocyanide,  resorcinol 
and  a-naphthol  tests. 

Formic  Acid. — Filter  the  residue  from  the  distillation,  con- 
centrate the  filtrate  to  a  few  cc.  by  evaporation  and  divide  into 
two  parts  for  the  following  reduction  tests : 

(a)  Reduction  of  Mercuric  to  Mercurous  Chloride. — Add 
a  few  drops  of  mercuric  chloride  solution  and  warm.  Formic 
acid,  if  present,  will  produce  a  white  precipitate  of  mercurous 
chloride  (calomel) : 

Mg(00CH)2  +  4HgCl2  =  2Hg2Cl2  +  MgCU  +  2HCI  +  2CO2. 

(b)  Reduction  of  Silver  Nitrate. — Warmed  with  silver  nitrate 

1  Pharmaceutische Zeitung  fur Russland  33,  373,  und Zeitschrif t fur analytische 
Chemie,  37,  60  (1898). 


40  DETECTION   OF   POISONS 

solution,  formic  acid  and  its  salts  produce  a  black  precipitate  of 
metallic  silver: 

Mg(00CH)2  +  4AgN03  =  4Ag  +  Mg(N03)2  +  2HNO3  +  2CO2. 
Detection  of  Chloral  Hydrate  in  Powders  or  Solutions 

Extract  a  powder  with  cold  water  containing  sulphuric  acid, 
filter,  extract  the  filtrate  several  times  with  ether  and  spon- 
taneously evaporate  the  ether  extracts  in  a  shallow  dish  or  on  a 
clock  glass.  Chloral  hydrate  imparts  to  the  residue  its  char- 
acteristic pungent  odor.  The  odor  of  chloroform  is  easily 
recognized  by  warming  the  residue  with  sodium  hydroxide 
solution : 

CCl3-CH(OH)2  +  KOH  =  CHCI3  +  H.COOK  +  H2O 

The  phenylisocyanide,  resorcinol  and  naphthol  tests,  as  well 
as  that  with  Nessler's  reagent,  should  be  applied  to  the  residue. 

In  the  case  of  an  aqueous  solution  of  chloral  hydrate,  first 
acidify  with  dilute  sulphuric  acid  and  repeatedly  extract  with 
ether.  Evaporate  the  ether  extracts  and  examine  the  residue  as 
already  described. 

Note. — Pure  chloral  hydrate  forms  transparent  crystals  which  are  dry,  perma- 
nent and  colorless.  This  compound  has  a  pungent  odor,  its  taste  being  caustic 
and  faintly  bitter.  It  dissolves  with  ease  in  water,  alcohol  and  ether;  and  in  5 
parts  of  chloroform.     It  melts  at  58°. 

Action  and  Fate  of  Chloral  Hydrate  in  the  Hiunan  Organism 

Applied  locally  chloral  hydrate  acts  as  a  strong  stimulant.  Taken  internally 
it  frequently  stimulates  the  stomach.  When  it  reaches  the  blood,  it  acts  like 
chloroform  in  paralyzing  the  brain,  spinal  cord  and  heart  but  usually  no  previous 
stimulation  is  noticeable.  There  is  marked  decrease  in  blood-pressure  due  to 
paralysis  of  the  blood-vessels.  Death  from  chloral  hydrate  poisoning  is  occa- 
sioned by  impaired  circulation  and  respiration,  in  consequence  of  which  the  quan- 
tity of  oxygen  taken  in  and  of  carbon  dioxide  given  off  is  considerably  diminished. 
H.  Meyer  has  shown  that  the  narcotic  action  of  chloral  hydrate  depends,  as  does 
that  of  all  compounds  of  the  alcohol  and  chloroform  group,  upon  the  affinity  of 
the  poison  for  lipoids,  the  fatty  constituents  of  the  nervous  system.  It  is  also 
held  by  the  blood,  especially  by  the  red  blood-corpuscles.  Later  it  appears 
unchanged,  most  abundantly  in  the  cells  of  the  brain  and  spinal  cord  (Kobert, 
"  Intoxikationen  ") . 

Only  very  little  chloral  hydrate  taken  internally  passes  as  such  into  the  urine. 
As  shown  by  v.  Mering  and  Musculus,^  the  greater  part  by  conjugation  with  gly- 

^Berichte  der  Deutschen  chemischen  Gesellschaft  8,  662  (1875);  and  v.  Mer- 
ing, Ibid.,  15,  1019  (1882). 


VOLATILE   POISONS  41 

curonic  acid  forms  urochloralic  acid  (CglliiClaO?)  which  is  eliminated  as  such  in 
the  urine.  This  conjugated  acid  undergoes  hydrolysis,  when  boiled  with  dilute 
acids,  and  gives  trichlor-ethyl  alcohol  and  free  dextro-rotatory  glycuronic  acid: 

CsHuCIaO;  +  H2O  =  CCI3-CH2OH  +  II00C-(CPI.0H)4-CH0 

Urochloralic  Trichlor-  Glycuronic 

acid.  ethyl  alcohol.  acid. 

Urochloralic  acid  is  therefore  trichlor-ethyl  glycuronic  acid.  It  is  crystalline 
and  with  heat  reduces  silver  solution  as  well  as  alkaline  copper  and  bismuth  so- 
lutions. Consequently  chloral  urine  behaves  much  like  sugar  urine  but  differs 
from  the  latter  in  being  strongly  laevo-rotatory.  The  reduction  of  the  aldehyde 
chloral,  to  its  corresponding  primary  alcohol,  trichlor-ethyl  alcohol,  is  especially 
noteworthy  as  regards  the  behavior  of  chloral  hydrate  in  the  human  organism. 

Quantitative  Estimation  of  Chloral  Hydrate  in  Blood  and  Tissues 

(ArchangelskyO 

Distil  the  material  for  12-20  hours  with  its  own  weight  of  20  per  cent,  phos- 
phoric acid,  repeating  the  process  if  the  distillate  is  turbid  or  yellow.  To  com- 
plete the  decomposition  of  chloral  hydrate  into  chloroform  and  formic  acid,  add 
50  cc.  of  sodium  hydroxide  solution  to  the  distillate  and  concentrate  on  the  water 
bath  to  about  20  cc.  Neutralize  the  solution  exactly  and  heat  for  6  hours  on  the 
water  bath  with  an  excess  of  mercuric  chloride  solution.  Finally  weigh  the 
precipitated  mercurous  chloride.  Satisfactory  results  were  obtained  by  this 
method  when  known  quantities  of  chloral  hydrate  were  added  to  blood  and 
organs.  Using  this  method  Archangelsky  has  shown  that  chloral  hydrate  is  not 
uniformly  distributed  in  the  blood  but  is  contained  especially  in  the  blood-cor- 
puscles. When  narcosis  begins  there  is  less  chloral  hydrate  in  the  brain  than  in 
the  blood.  But  later  the  percentage  of  the  poison  in  the  brain  is  higher  than  in 
the  blood.  Archangelsky  has  further  shown  how  much  chloral  hydrate  the  blood 
must  contain  before  narcosis  can  appear.  A  dog's  blood  must  contain  0.03-0.05 
per  cent.     When  the  blood  contains  o.i  2  per  cent.,  respiration  ceases. 

IODOFORM 

Iodoform  crystallizes  in  shining  hexagonal  leaflets  or  plates.     It  may  also 

T  appear  as  a  rather  fine  crystalline  powder,  lemon-yellow  in  color  and 

I  having  a  penetrating  odor  somewhat  like  saffron.     The  melting-point 

I — C — I    of  iodoform  is  approximately  120°.     It  is  nearly  insoluble  in  water; 

soluble  in  50  parts  of  cold  and  in  about  10  parts  of  boiling  alcohol; 

and  soluble  in  6  parts  of  ether.     It  is  also  freely  soluble  in  chloroform. 


H 


Detection  of  Iodoform 

Iodoform  distils  quite  easily  with  steam  and  gives  a  milky- 
distillate  having  a  characteristic  odor.  Extract  this  distillate 
with  ether  and  carefully  test  the  residue  left  by  the  spontaneous 

^Archiv  fur  experimenteUe  Pathologic  und  Pharmakologie,  46,  347  (1901). 


42  DETECTION   OF  POISONS 

evaporation  of  the  solvent.  If  much  iodoform  is  present, 
it  will  form  yellow  hexagonal  plates.  Dissolve  the  ether 
residue  in  a  little  alcohol,  and  use  this  solution  for  the  following 
tests : 

I.  Lustgarten's^  Test. — Gently  warm  a  few  drops  of  alcoholic 
iodoform  solution  in  a  test-tube  with  a  little  sodium  phenolate 
(CeHs.ONa)  solution.  2  If  iodoform  is  present,  a  red  substance 
will  be  deposited  on  the  bottom  of  the  tube.  A  few  drops 
of  dilute  alcohol  will  dissolve  this  precipitate  with  a  carmine- 
red  color. 

Also  make  the  resorcinol  and  phenylisocyanide  tests  (see 
page  36). 

NITROBENZENE 

Nitrobenzene  has  a  strong  poisonous  action.     Administration  of  very  small 
quantities  of  this  compound  has  produced  death  in  human  beings.     There  are 
■jv^Q         records  in  the  literature  of  several  cases  where  20  drops,  and  even 
I  7  to  8  drops,  have  caused  fatal  results.     But  on  the  other  hand 

C  complete  recovery  has  followed  poisoning  by  much  larger  doses. 

xrr^    r XT    ^^^^  poisonings  have  come  also  from  inhaling  nitrobenzene  vapor. 
I  11        Within  recent  years  nitrobenzene  has  been  used  to  some  extent  as 

HC        CH    an  abortifacient.     Nitrobenzene  poisons  the  blood  and  changes  its 
\/'         appearance.     The  blood  has  a  chocolate  color  and  at  the  same  time 
^  the  red  blood-corpuscles  change  their  shape  and  go  into  solution. 

As  a  result  the  blood  is  incapable  of  uniting  with  oxygen.  The 
blood  of  persons  poisoned  by  nitro  benzene  is  said  to  contain  less  than  i  per 
cent,  of  oxygen  so  that  death  is  caused  by  asphjotiation.  Healthy  blood 
contains  about  17  per  cent,  of  oxygen  by  volume.  There  seems  to  be  no 
methaemoglobin  in  blood  containing  nitrobenzene.  Such  blood  examined 
spectroscopicaUy  shows  the  two  oxyhsemoglobin  bands  and  also  a  special 
absorption-band  between  C  and  D  (Fihlene's  nitrobenzene  band).  It  is  proba- 
ble that  the  slight  solubility  .of  this  poison  necessitates  a  definite  incubation 
period,  for  2  to  3  hours  usually  elapse  after  nitrobenzene  has  been  taken  before 
signs  of  intoxication  appear.  A  woman,  who  had  taken  10  drops  of  mirbane  oil 
as  an  abortifacient,  gave  no  indication  of  intoxication,  that  is  to  say,  uncon- 
sciousness and  cyanosis,  for  8  hours  after  taking  the  poison. 

Nitrobenzene  not  only  profoundly  changes  the  blood  but  it  irritates  and 
paralyzes  the  central  nervous  system  (see  R.  Robert,  "Intoxikationen"). 

Some  nitrobenzene  passes  into  the  urine  but  the  organism  does  not  appear  to 
convert  it  into  aniline.     In  nitrobenzene  poisoning  human  urine  contains  a 

1  Monatshefte  fiir  Chemie,  3,  715  (1882). 

*  Prepare  sodium  phenolate  solution  by  mixing  20  grams  of  phenol  with  40 
grams  of  sodium  hydroxide  and  70  grams  of  water. 


VOLATILE    POISONS  43 

brown  pigment  but  only  rarely  hcemoglobin  or  mcthsemoglobin.  Urine  contain- 
ing nitrobenzene  will  reduce  Fehling's  solution.  It  is  also  unfermcntable  and  dis- 
tinctly Isevo-rotatory.  A  conjugated  glycuronic  acid  is  possibly  concerned  in 
these  reactions. 

Detection  of  Nitrobenzene 

In  nitrobenzene  poisoning  the  urine  and  all  the  organs  have 
the  odor  of  this  compound.  For  the  chemical  tests  the  material 
should  first  be  distilled  with  water.  Nitrobenzene  distils  quite 
easily  with  steam  and  appears  in  the  distillate  as  yellowish 
globules.  These  are  heavier  than  water  and  have  a  character- 
istic odor.  Vigorously  agitate  the  globules,  when  separated  as 
completely  as  possible  from  water,  with  granulated  tin  and  a 
few  cc.  of  concentrated  hydrochloric  acid,  until  there  is  no  odor 
of  nitrobenzene.  Pour  the  acid  solution  from  undissolved  tin, 
and  add  an  excess  of  potassium  hydroxide  solution  to  decompose 
the  double  chloride  of  aniline  and  tin.  Extract  free  aniline 
with  ether.  Withdraw  the  aqueous  Hquid  from  the  separating 
funnel,  and  evaporate  the  ether  extract  spontaneously  in  a 
small  glass  dish.  Aniline,  formed  by  reducing  nitrobenzene, 
will  remain  as  globules  which  usually  have  a  red  or  brown  color. 
Dissolve  these  globules  by  agitation  with  water,  and  use  this 
solution  for  the  hypochlorite  and  phenylisocyanide  tests  (see 
pages  45  and  36). 

Mechanism  of  the  Reaction. — Nitrobenzene  is  reduced  by  nascent  hydrogen 
to  aniline  (a)  which  combines  with  the  excess  of  hydrochloric  acid  forming  aniline 
hydrochloride  (j8).  From  the  latter  compound  potassium  hydroxide  liberates 
aniline  (7) : 

(a)   CeHe-NOa  +  6H        =  CeHs-NHz  +  2H2O, 

(^)    CeHs-NHo  +  HCl     =  CeHe-NHa.HClS 

(t)    CeHeNHz.HCl  +  KOH  =  CeHs-NHa  +  H2O  +  KCl. 

^  Organic  ammonium  bases  resemble  ammonia  in  combining  with  acids  to  form 
salts.  Trivalent  nitrogen  of  the  free  base  is  changed  to  pentavalent  nitrogen  in 
the  salt: 

III  ,H  V  /| 

CsHb  =  N^      +  HCl  =  CsHs  -  N^g 

^  ^Cl 

Aniline  Aniline  h3-drochloride 


44  DETECTION   OF  POISONS 

ANILINE 

Toxic  Action. — Aniline  is  moderately  toxic  in  its  action.     Doses  of  1.5  to  2 
grams,  administered  in  the  course  of  a  day,  have  proved  fatal  to  small  dogs.     It 
is  not  possible  to  state  definitely  the  average  lethal  dose  for  human  beings.     Very 
NH        serious  results  are  said  to  have  followed  a  dose  of  3  or  4  grams  of 
I  aniline.     The  lethal  dose  is  certainly  less  than  25  grams,  for  that 

C  quantity  of  aniline  was  sufi&cient  to  kill  a  healthy  man.     Even 

j^\  inhalation  of  aniline  vapor  may  cause  severe,  or  fatal  intoxications. 

I        II  Aniline   produces   methaemoglobin   and   therefore   poisons    the 

HC      CH      blood.     The  conversion  of  oxyhsemoglobin  into  methaemoglobin 
\/  by  aniline  may  be  demonstrated  by  adding  an  aqueous  aniline 

^  solution  to  blood  in  a  test-tube.     Aniline  changes  their  form  and 

partially  decomposes  red  blood-corpuscles.  Thereby  the  quan- 
tity of  available  oxygen  in  the  blood  is  so  diminished  that  it  amounts  to  only  5 
to  10  volumes  instead  of  15  to  20,  the  normal  quantity.  The  number  of  red 
blood-corpuscles  is  diminished  in  aniline  poisoning  but  not  that  of  the  white  blood- 
cells. 

R.  V.  Engelhardt  has  shown  that  aniline  is  partly  changed  in  the  human  organ- 
ism into  aniline  black,  or  into  a  similar  compound  insoluble  in  water.  At  the 
climax  of  aniline  poisoning  blue-black  granules  may  be  seen  in  every  drop  of  blood 
and  also  in  the  urine.  Aniline  is  oxidized  in  the  system  to  para-aminophenol 
(C6H4.0H,NH2(i,4))  .  Like  all  phenols  this  compound  forms  an  ethereal  sul- 
phate with  sulphuric  acid,^  namely,  para-aminophenyl-sulphuric  acid  (HO.SO2.- 
O.C6H4.NH2(i,4).  This  acid  is  eliminated  through  the  kidneys  as  an  alkali  salt 
and  then  appears  in  the  urine  A  part  of  the  para-aminophenol  is  also  elim- 
inated as  a  conjugate  of  glycuronic  acid.^ 

The  reduction  of  Fehling's  solution  by  urine  containing  aniline  is  due  to 
this  conjugated  acid.  In  severe  cases  of  poisoning  unchanged  aniline  has  also 
been  found  in  the  urine.  Usually  urine  that  contains  aniline  has  a  very  dark 
color.  Besides  the  substances  mentioned,  a  dark  pigment  has  been  detected 
in  urine  in  aniline  poisoning  as  well  as  haemoglobin,  methaemoglobin  and  an 
abundance  of  urobilin  (R.  Kobert,  "Intoxikationen")  • 

Detection  of  Aniline 

Aniline  is  a  rather  feeble  base  and  part  of  it  will  pass  over 
with  steam,  when  the  acid  solution  is  distilled.  There  will  be 
enough  in  the  distillate  for  detection  by  the  tests  described  be- 

^  This  conjugation  takes  place  with  elimination  of  water: 

H2N.C6H4.0H  +  HO.S02.0H  =  H2O-I-H2N.C6H4.O.SO2.OH  (i,  4) 

2  Glycuronic  acid,  C6Hio07=Q_^C-(CH.OH)4-COOH,    is   closely    related    to 

glucose.  It  is  an  uncrystallizable  syrup.  If  its  aqueous  solution  is  boiled,  the 
acid  is  partly  converted  into  the  internal  anhydride,  glycurone  (CeHsOe),  which 
crystallizes  well. 


VOLATILE    POISONS  45 

low.  In  estimating  aniline  quantitatively  in  any  kind  of  mate- 
rial the  distillation  must  be  as  complete  as  possible.  Mix  the 
substance  with  water,  make  strongly  alkaline  with  sodium 
hydroxide  or  carbonate  solution  and  distil  in  a  current  of  steam. 
Since  30  parts  of  water  at  15°  dissolve  i  part  of  anihne,  the 
distillate  may  contain  considerable  of  this  amine.  When  the 
quantity  is  large,  oil-drops  will  appear.  An  aqueous  aniline 
solution  (aniline  water)  colors  pine  wood  and  elder  pith 
intensely  yellow.  The  following  tests  should  be  used  for 
aniline : 

1.  H3rpochlorite  Test. — Add  a  few  drops  of  aqueous  calcium  or 
sodium  hypochlorite  solution  drop  by  drop  to  a  portion  of  the 
distillate.  A  violet-blue  or  purple- violet  color,  gradually  chang- 
ing to  a  dirty  red,  will  appear  if  anihne  is  present.  Addition  of  a 
little  dilute  aqueous  phenol  solution  containing  some  ammonia 
will  produce  a  blue  color  which  is  quite  stable.  This  test  is 
sensitive  i  : 66,000.^ 

2.  Phenylisocyanide  Test. — Heat  a  portion  of  the  distillate 
with  a  few  drops  of  chloroform  and  potassium  hydroxide  solu- 
tion. The  repulsive  odor  of  phenylisocyanide  will  show  the 
presence  of  aniline. 

3.  Bromine  Water  Test. — Bromine  water  added  to  a  solution 
containing  aniline  will  produce  a  flesh-colored  precipitate. 
This  test  is  sensitive  i :  66,000. 

4.  Chromic  Acid  Test.^ — Mix  a  trace  of  pure  aniline  with  4  to 
5  drops  of  concentrated  sulphuric  acid  in  a  porecelain  dish  and 
add  a  drop  of  aqueous  potassium  dichromate  solution.  After 
a  few  minutes  the  mixture  beginning  at  the  edge  will  take  on  a 
pure  blue  color.  Addition  of  1-2  drops  of  water  produces  at 
once  a  deep  blue  color.  To  apply  this  test  to  the  distillate, 
first  extract  with  ether,  evaporate  the  ether  solution  and  test  an 
oily  residue  as  described. 

^  Test  this  experimentally  with  very  little  aniline.  For  example,  dissolve  a 
small  drop  in  30  cc.  of  water  and  take  only  2-3  cc.  of  this  dilute  solution  for  the 
test. 

2  Beissenhirtz  reaction,  Annalen  der  Chemie  und  Pharmazie,  87..  376  (1853). 


46  DETECTION   OF  POISONS 

CARBON  BISULPHIDE 

Carbon  disulphide,  CS2,  is  a  colorless  liquid  having  a  characteristic  odor  and  a 
high  index  of  refraction.     It  is  only  slightly  soluble  in  water.     There  is  some 
difference  of  opinion  as  regards  the  solubility  of  carbon  disulphide  in  water. 
1000  cc.  of  water  dissolve 

13-14°  2.03  parts  (Page) 

15-16°  1. 81  parts  (Chancel;  Parmentier) 

15-16°  2-3  parts  (Ckindi) 

15-16°  3.5-4.52  parts  (Peligot) 

Carbon  disulphide  is  miscible  in  aU  proportions  with  absolute  alcohol,  ether, 
ethereal  and  fatty  oils. 

Toxic  Action. — Carbon  disulphide  administered  internally  has  a  very  poisonous 
action  upon  the  blood  causing  especially  decomposition  of  red  blood-corpuscles. 
Even  inhalation  of  carbon  disulphide  vapor  frequently  occasions  severe  poisoning. 
Carbon  disulphide  was  formerly  considered  a  typical  producer  of  methaemoglobin 
but  recent  investigations  have  not  confirmed  this  opinion.  It  has  a  very  injurious 
action  upon  the  red  blood-corpuscles  and  causes  haemolysis.  R.  Kobert  (Intox- 
ikationen)  states  that  its  power  of  dissolving  lipoids  is  responsible  for  its  injuri- 
ous action  upon  the  blood  and  the  central  nervous  system.  E.  Harmsen^  has 
recently  come  to  practically  the  same  conclusion.  He  considers  carbon  disul- 
phide a  powerful  blood  poison  because  it  decreases  the  number  of  red  blood-cor- 
puscles and  the  quantity  of  hsemoglobin  and  brings  about  a  leucocytosis.'^ 

Detection  of  Carbon  Disulphide 

Carbon  disulphide  distils  very  slowly  with  steam.  Con- 
sequently the  second  or  third  fraction  of  the  distillate  should 
be  used  in  testing  for  this  substance.  If  40  cc.  are  distilled 
from  100  cc.  of  water  containing  2  drops  of  carbon  disulphide, 
the  following  10  cc.  will  give  a  distinct  test.  If  the  quantity 
of  carbon  disulphide  is  small,  it  will  remain  in  solution.  Such 
a  solution  does  not  have  a  strong  odor.  Carbon  disulphide  may 
be  recognized  by  the  following  tests: 

I.  Lead  Acetate  Test. — ^Add  a  few  drops  of  lead  acetate 
solution  to  the  liquid  containing  carbon  disulphide.  It  will 
cause  neither  a  precipitate  (distinction  between  CS2  and  H2S) 
nor  a  color.  Add  excess  of  potassium  hydroxide  solution  and 
boil.  A  black  precipitate  (PbS)  will  appear.  This  is  a  very 
delicate  test. 

1  Vierteljahrsschrift  fiir  gerichtUche  Medizin,  30,  422  (1905). 

^  Leucocytosis  means  a  temporary  increase  in  the  number  of  white  blood- 
corpuscles  (leucocytes)  as  compared  with  the  number  of  red  blood-corpuscles. 
Normally  there  are  about  350  red  to  i  white  blood-corpuscle,  whereas  in  Icucocy- 
tosis  the  proportion  is  20  :  i. 


VOLATILE   POISONS  47 

2.  Sulphocyanate  Test. — Heat  an  aqueous  solution  of  carbon 
disulphide  for  a  few  minutes  with  concentrated  ammonium 
hydroxide  solution  and  alcohol.  Ammonium  sulphocyanate 
(H4NSCN)  is  formed  together  with  ammonium  sulphide. 
Concentrate  this  solution  upon  the  water-bath  to  about  i  cc. 
and  acidify  with  dilute  hydrochloric  acid.  Add  a  drop  of 
ferric  chloride  solution  and  a  deep  red  color  will  appear.  This 
test  will  show  even  traces  of  carbon  disulphide,  for  example 
0.05  gram  in  i  cc.  of  water. 

Mechanism  of  the  Reaction: 

(a)  4NH3  +  CS2  =  (H4N)SCN  +  (H4N)2S, 

(13)   FeCls  +  3(H4N)SCN      =  (Fe(SCN)3  +  3(H4N)C1. 

3.  Xanthogenate  Test. — Shake  a  few  cc.  of  distillate  for 
several  minutes  with  3  or  4  times  its  volume  of  saturated  solu- 
tion of  potassium  hydroxide  in  absolute  alcohol.  Faintly 
acidify  the  solution  with  acetic  acid  and  add  i  or  2  drops  of 
copper  sulphate  solution.  If  carbon  disulphide  is  present,  a 
brownish  black  precipitate  of  cupric  xanthogenate  will  appear. 
This  will  soon  change  to  a  yellow,  flocculent  precipitate  of 
cuprous  xanthogenate,  CS(SCu)  (OC2H5) .  Vitali's  procedure  is 
somewhat  different  and  consists  in  adding  soHd  ammonium 
molybdate  to  the  alkaUne  reaction-product  and  then  in  acidify- 
ing with  dilute  sulphuric  acid.  The  appearance  of  a  red  color 
indicates  carbon  disulphide. 

Mechanism  of  the  Reaction. — Alcoholic  potassium  hydroxide  acts  like 
potassium  alcoholate  (C2H6-OK)  and  converts  carbon  disulphide  into  potassium 
xanthogenate 

/SK 
CS2  +  C2H6-OK  =  C==S 

\OC2H6 
This  compound  treated  with  cupric  salts  gives  first  a  brownish  black  precipitate 
of  cupric  xanthogenate : 

/SK  /S— 

2C=S  +  CUSO4  =  (S  =  C<  )2Cu  -f-  K2SO4 

\OC2H6  \OC2H6 

The  cupric  salt  then  forms  cuprous  xanthogenate  and  ethyl  xanthogen  disulphide : 
/OC2HB  /OC2H6  /OC2H5 

S  =  C<  S  =  C<  S  =  C< 

\S\  ^S  ^S  -  Cu 

2  ^Cu     =  I        +  I 

/S^  /S  /S  —  Cu 

s  =  c<  s  =  c<  s  =  c< 

\OC2H5  ^OC2H6  ^OCoHs 

Cupric  Ethyl  xanthogen  Cuprous 

xanthogenate  disulphide  xanthogenate 


48  DETECTION   OF  POISONS 

Quantitative  Estimation  of  Carbon  Bisulphide  in  Air 

Inhalation  of  air  containing  carbon  disulphide  has  frequently  given  rise  to 
chronic  poisoning.  Persons  thus  affected  have  usually  been  laborers  in  rubber  fac- 
tories. Consequently  experiments  have  been  made  to  determine  the  maximum 
quantity  of  carbon  disulphide  air  may  contain  without  injury  to  health.  The 
results  of  these  investigations  may  be  summarized  as  follows : 


CS2in 

mgrs. 

Result 

per  liter  of  air 

I. 

0.5-' 

0.8 

No  injurious  effect. 

2. 

1-3 

Slight  uneasiness  after  several 
hours. 

3- 

3-4 

Uneasiness  in  30  minutes. 

4- 

6.0 

Uneasiness  in  20  minutes. 

5- 

ID  .  0 

Paralysis  attended  by  after-effects  last- 
ing several  days. 

The  exact  danger  Umit  for  persons  obliged  to  live  for  weeks  at  a  time  in  an  atmo- 
sphere containing  carbon  disulphide  should  be  placed  below  3  mg.  per  liter 
of  air.  Air  in  factories,  where  operatives  work  in  presence  of  carbon  disulphide 
vapor,  should  never  exceed  this  limit.  In  rubber  factories  the  air  is  said  fre- 
quently to  contain  2.5  to  3  mg.  per  liter.  Since  experiments  have  shown 
that  93  to  96  per  cent,  of  the  carbon  disulphide  breathed  was  exhaled  unchanged, 
an  exceedingly  small  quantity  is  capable  of  producing  toxic  symptoms. 

Procedure. — Place  a  saturated  alcoholic  solution  of  potassium  hydroxide  in  a 
P^Hgot  absorption-tube  and  draw  through  this  solution  10  to  20  liters  of  air  con- 
taining carbon  disulphide  vapor.  A  quantitative  formation  of  potassium  xan- 
thogenate  (see  above)  will  take  place. 

Dilute  the  contents  of  the  receiver  at  the  end  of  the  experiment  with  96  per 
cent,  alcohol  and  bring  the  volume  to  50  cc.  Measure  an  aliquot  portion  of  this 
solution  and  dilute  with  water.  Faintly  acidify  the  solution  with  acetic  acid 
and  remove  excess  of  acid  with  acid  sodium  carbonate.  Add  freshly  prepared 
starch  solution  and  o.  i  n-iodine  solution  until  there  is  a  permanent  blue  color. 

Iodine  converts  potassium  xanthogenate  according  to  equation  (I)  into  ethyl 
xanthogen-disulphide : 

KS.CS.OC2H6  S.CS.OC2H6 

I.  I2  -}-  =  2KI  -1-  I 

KS.CS.OC2H2  S.CS.OC2H6 

E.  Rupp  and  L.  Krauss^  think  the  action  of  iodine  upon  potassium  xanthogen- 
ate is  expressed  by  equation  (II) : 

II.  2KS.CS.OC2H5  -1-  H2O  -f  2I  =  KS.CS.SK  +  2C2H6.OH  -t-  2HI  -f  S. 
Both  equations  require  the  same  quantity  of  iodine,  namely,  2  atoms  for  2 
molecules  of  xanthogenate.     A  difference  therefore  in  the  mechanism  of  the 
reaction  has  no  influence  on  the  combining  relations  of  the  iodine  and  the  method 
is  apphcable  to  the  quantitative  determination  of  xanthogenate. 

1000  cc.  of  0.1  n-iodine  solution,  containing  o.i  gram-atom  of  iodine,  corre- 
spond to  0.1  gram-molecule  of  CS2   =  7.6  grams. 

^  Berichte  der  Deutschen  chemischen  Gesellschaft  35,  4257  (1902). 


VOLATILE   POISONS  49 


ETHYL  ALCOHOL 


Fate  in  the  Human  Organism. — Alcohol  brought  in  contact  with  many  dffer- 

ent  parts  of  the  organism  is  very  rapidly  absorljcd,  but  especially  easily  from  an 

empty  stomach.     Although  there  is  practically  no  absorption  of  non-volatile 

TT  aqueous  liquids   from  the  stomach,   alcohol  is  freely  absorbed. 

I  After  absorption  it  passes  into  the  blood  and  is  then  distributed 

H — C — H      to  all  organs  (see  chloral  hydrate).     Experiments  upon  dogs,  colts 

I  and  adult  horses  (see   Kobert,   "Intoxikationen")    have   shown 

I  that  blood  at  the  climax  of  narcosis  contains  0.72  per  cent,  of 

jH  alcohol.     There  is  stupor  even  when  C.I 2  per  cent,  is  present. 

There  is  difference  of  opinion  among  toxicologists  regarding 
alcoholic  intoxication,  as  to  whether  the  poison  is  distributed  uniformly  through- 
out the  body,  or  accumulated  in  the  brain  in  larger  quantity  than  in  other 
organs.  The  following  percentages  of  alcohol,  found  in  the  organs  of  a  man, 
who  had  died  at  the  climax  of  severe  acute  alcohol  poisoning,  lend  support  to 
the  latter  view:  liver  0.21,  brain  0.47  and  blood  0.33  per  cent.  It  appears  from 
these  results  that  the  brain  takes  up  an  especially  large  quantity  of  alcohol. 

Uncertainty  concerning  the  subsequent  fate  of  alcohol  in  the  organism  has 
finally  been  removed.  Experiments  have  shown  that  alcohol  is  never  eUminated 
unchanged  through  the  skin.  At  most  only  1-1.5  per  cent,  passes  o£f  through 
the  kidneys  and  only  i  .6-2  per  cent,  throught  the  lungs.  Strassmann^  found  the 
quantity  eliminated  by  the  lungs  somewhat  higher  (5-6  per  cent.)  and  by  the 
kidneys  1-2.5  P^r  cent.  The  remainder  is  completely  oxidized  in  the  human 
organism  to  carbon  dioxide  and  water. 

B.  Fischer  found  the  following  quantities  of  alcohol  in  organs  removed  from  a 
man  who  had  probably  died  from  drinking  too  much  brandy: 

Weight                              Organ  Alcohol 

2720  grams  Stomach  and  intestines  30.6    grams 

2070  grams  Heart,  lungs  and  blood  10.85  grams 

1820  grams  Kidneys  and  liver  7.8    grams 

1365  grams  Brain  4.8    grams 

Detection  of  Ethyl  Alcohol 

Ethyl  alcohol  distils  easily  with  steam  and  consequently 
most  of  it  will  be  in  the  first  fraction.  If  present  in  sufficient 
quantity,  it  can  be  recognized  in  the  distillate  by  its  odor.  The 
following  tests  should  be  made: 

I.  Lieben's  Iodoform  Test.^ — Gently  warm  the  liquid  (40- 
50°),  add  a  few  cc.  of  aqueous  iodo-potassium  iodide  solution,  or 
a  small  crystal  of  iodine,  and  enough  potassium  hydroxide 
solution  to  give  the  liquid  a  distinct  yellow  to  brownish  color. 

^  Pfluger's  Archiv,  49,  315  (1891). 

2  Annalen  der  Chemie  und  Pharmazie,  Supplement  Band,  7,  21S. 
4 


50 


DETECTION   OE   POISONS 


If  alcohol  is  present ,  a  yellowish  white  to  lemon-yellow  precipi- 
tate of  iodoform  will  appear  immediately,  or  as  the  solution 
cools.     If  the  quantity  of  alcohol  is  very  small,  a  precipitate  will 

form  on  long  standing.  When  iodo- 
form is  deposited  slowly,  the  crystals 
are  very  perfect  hexagonal  plates  and 
stars  (see  Fig.  9). 


Fig.  9. — Iodoform  Crystals. 


Note. — This  iodoform  test  is  very  delicate 
but  not  characteristic  of  ethyl  alcohol,  since 
other  primary  alcohols,  except  methyl  alcohol, 
and  many  secondary  alcohols,  as  weU  as  their 
oxidation  products,  aldehydes  and  ketones, 
give  iodoform  under  the  same  conditions. 
Acetic  ether,  aceto-acetic  ether,  lactic  acid,  etc., 
also  give  iodoform. 

The  correct  explanation  of  the  iodoform  reaction  is  probably  the  following: 
Iodine  and  potassium  hydroxide  form  potassium  hypo-iodite  (KOI)  by  reaction 
(a).  This  compound  brings  about  the  oxidation  of  alcohol  to  acetic  aldehyde 
(jS)  and  at  the  same  time  substitutes  iodine  for  hydrogen  in  the  latter  (7).  Finally 
tri-iodo-acetic  aldehyde  is  decomposed  by  the  excess  of  potassium  hydroxide  into 
iodoform  and  potassium  formate  (5) : 

(a)   2KOH  +  I2         =  KI  +  H2O  +  KOI, 

()3)    CH3.CH2.OH  +  KOI    =  CH3.CHO  +  H2O  +  KI, 
(7)    CH3.CHO        +  3KOI  =  3KOH         +  CI3.CHO, 
(5)    CI3.CHO         +  KOH  =  CHI3  +  H.COOK. 

2.  Berthelot's  Test. — Shake  the  liquid  containing  alcohol 
with  a  few  drops  of  benzoyl  chloride  and  about  5  cc.  of  sodium 
hydroxide  solution  (10  per  cent.),  until  the  irritating  odor  of 
benzoyl  chloride  has  gone.  The  aromatic  odor  of  ethyl  ben- 
zoate  will  appear. 

CeHs.COCl  +  C2H5.OH  +  KOH  =  C6H6.CO.OC2H6  +  KCl  +  H2O 

Ten  cc.  of  0.5  per  cent,  alcohol  will  give  a  distinct  odor  of  this 
ester. 

3.  Chromic  Acid  Test. — Warm  the  hquid  containing  alcohol 
with  dilute  sulphuric,  or  hydrochloric  acid,  and  add  i  or  2  drops 
of  very  dilute  potassium  dichromate  solution.  The  color  of  the 
liquid  will  change  from  red  to  green,  and  at  the  same  time  the 
odor  of  acetaldehyde  will  be  recognized.  This  test  is  not  char- 
acteristic of  alcohol,  because  many  other  volatile  organic 
compounds  behave  similarly. 


VOLATILE    POIf?ONS  51 

Mechanism  of  the  Reaction 

(a)  KaCrjO,  +     H2SO4  =  K2SO4  +  HiCr-^OjCHjO  +  aCrOa), 

(13)  3  C2H5.OH  +  2  Cr03  +  3  H2SO4  =  3  CH3.CHO  +  Cr2(S04)3+  6H2O. 

Acetaldehyde 

4.  Ethyl  Acetate  Test. — Mix  the  liquid  containing  alcohol 
with  the  same  volume  of  concentrated  sulphuric  acid.  Add  a 
very  small  quantity  of  anhydrous  sodium  acetate  and  heat. 
Ethyl  acetate  will  be  recognized  by  its  odor. 

(a)   C2H6.OH  +  H2SO4  =  C2H6O.SO2.OHI  +  H2O, 

(;8)   CH3.CO.ONa  +  C2HBO.SO2.OH  =  CH3.CO.OC2H6  +  NaHSO*. 

5.  Vitali's  Test.— Thoroughly  mix  a  few  cc.  of  distillate  in  a 
glass  dish  with  a  small  piece  of  solid  potassium  hydroxide  and 
2  or  3  drops  of  carbon  disulphide.  Let  this  mixture  stand  for  a 
short  time  without  warming.  When  most  of  the  carbon  disul- 
phide has  evaporated,  add  a  drop  of  ammonium  molybdate 
solution  and  then  an  excess  of  dilute  sulphuric  acid.  If  the  dis- 
tillate contains  alcohol,  a  red  color  will  appear.  Potassium 
xanthogenate  (CS(OC2H5)(SK))  is  first  formed.  This  com- 
pound gives  a  red  color  with  ammonium  molybdate.  Acetone 
and  acetaldehyde  produce  a  similar  color.  This  test  is  given 
distinctly  by  5  per  cent,  aqueous  alcohol  solution. 

ACETONE 

Human  urine  almost  always  contains  a  very  small  quantity  of  acetone  as  a 

physiological  constituent.     Under  pathological  conditions,  especially  in  diabetes 

jj  meUitus  (diabetic  acetonuria),  urine  contains  much  more.     It  is 

I  also  present  in  urine  in  protracted  high  fever,  digestive  disturb- 

H     C     H       ances,  severe  forms  of  carcinoma  (carcinomatous  acetonuria),  etc. 

1 „        Finally,  acetone  has  been  found  in  urine  in  considerable  quantity 

I  in  various  intoxications  (toxic  acetonuria) ,  for  example,  in  poison- 

H — C — H      ing  by  phosphorus,  carbon  monoxide,  atropine,  curare,  antipyrine, 

I  pyrodine,    sulphuric    acid,    extract  of  male  fern;  in  chronic  lead 

poisoning;  and  in  chronic  morphinism  after  discontinuance  of  the 

drug   (see  R.   Kobert,   "Intoxikationen"). 

Acetone  is  not  poisonous  nor  in  the  least  corrosive.  Man  and  animals  can 
tolerate  considerable  quantities  of  acetone  taken  internally.  It  seems  to  produce 
no  effect,  though  it  may  possibly  possess  very  feeble  narcotic  properties.  Arch- 
angelsky  found  dogs  to  show  signs  of  narcosis  when  the  blood  contained  0.5  per 

O^      /OC2H6 

'  The  structural  formula  of  ethyl  sulphuric  acid  is      ^S<f 

O^     ^OH 


52  DETECTION   OF   POISONS 

cent,  of  acetone.  Even  smaller  doses  produce  narcosis  in  rabbits  and  have  an 
injurious  action  upon  the  blood  and  kidneys. 

Distillates  from  human  urine,  as  well  as  from  blood  and  various  organs,  as 
liver,  spleen,  kidneys,  brain,  etc.,  often  contain  acetone,  or  more  correctly  per- 
haps, substances  like  acetone.  This  is  especially  the  case  when  cadaveric  mate- 
rial has  begun  to  putrefy. 

Acetone  is  a  clear,  colorless  liquid  boiUng  at  56°.  It  has  a  pecuUar,  fruity  odor 
and  is  neutral  in  reaction.  It  is  miscible  in  all  proportions  with  water,  alcohol 
and  ether.     It  distils  easily  with  steam. 

Detection  of  Acetone 

1.  Lieben's  lodofonn  Test. — ^Add  a  few  cc.  of  aqueous  iodo- 
potassium  iodide  solution,  or  a  small  crystal  of  iodine,  to  an 
aqueous  solution  of  acetone  and  then  potassium  hydroxide 
solution  drop  by  drop  until  the  color  is  yellow.  Iodoform 
immediately  separates,  even  in  the  cold,  as  a  yellowish  white 
precipitate  which  is  usually  amorphous.  Acetone  differs  from 
alcohol  in  giving  iodoform,  when  ammonium  hydroxide  solution 
is  substituted  for  potassium  or  sodium  hydroxide  solution 
(Gunning's  acetone  test). 

Acetaldehyde  resembles  acetone  in  giving  iodoform  in  the  cold 
and  under  conditions  the  same  as  those  stated  above. 

Note. — Potassium  hypo-iodite  (a)  probably  converts  acetone  into  tri-iodo- 
acetone  (CH3.CO.CI3)  (|S)  and  this  compound  is  then  decomposed  by  potassium 
hydroxide  into  iodoform  and  potassium  acetate  (7) : 

(«)  6K0H  +  3I2       =  3KI  +  3KOI  +  3H2O, 

(/3)   CH3.CO.CH3  +  3KOI  =  CH3.CO.CI3    +  3KOH, 

(7)   CH3.CO.CI3    +  KOH  =  CHI3  -f  CH3.CO.OK, 

2.  Legal's  Test. — Add  a  few  drops  of  freshly  prepared  sodium 
nitroprusside  solution  to  a  liquid  containing  acetone,  and  then 
potassium  hydroxide  solution.  A  red  or  reddish  yellow  color 
will  appear.  This  color  soon  changes  to  yellow.  Add  an 
excess  of  acetic  acid  to  the  solution.  The  solution  will  now 
have  a  carmine  to  purpHsh  red  color,  according  to  the  quantity 
of  acetone  present.     Heat  will  change  this  color  to  violet. 

Alcohol  does  not  give  Legal's  test,  though  acetaldehyde  does.  The  red  color 
caused  by  aldehyde  fades  upon  addition  of  acetic  acid,  and  changes  to  green  with 
heat.  Le  Nobel  states  that  ammonium  hydroxide,  or  ammonium  carbonate  solu- 
tion, may  be  substituted  for  potassium  hydroxide  solution  in  Legal's  test,  but 
under  these  conditions  the  red  color  is  very  slow  to  appear.  Le  Nobel's  modifica- 
tion, however,  ehminates  the  possibility  of  confusing  acetone  with  acetaldehyde. 


VOLATILE    POISONS  Od 

3.  Penzoldt's  Test. — Prepare  a  hot,  saturated,  aqueous 
ortho-nitro-benzaldehyde  (C6H4.N02.CHO(i,2))  solution  and 
allow  it  to  cool.  Add  this  solution  to  the  liquid  containing 
acetone,  and  also  some  sodium  hydroxide  solution.  At  first 
the  color  of  the  mixture  is  yellow.  It  then  becomes  green,  and  a 
blue  precipitate  of  indigotin  is  formed  in  10  to  15  minutes. 
When  indigotin  is  present  in  traces  only,  shake  the  solution  with 
chloroform.  This  solvent  will  dissolve  the  coloring  matter  and 
become  blue. 

4.  Reynold's  Test — Acetone  will  dissolve  freshly  precipi- 
tated mercuric  oxide,  and  this  test  is  based  upon  this  property. 
Add  mercuric  chloride  solution  to  the  distillate,  and  an  alcoholic 
potassium  hydroxide  solution.  Shake  thoroughly  and  filter. 
Add  ammonium  sulphide  solution  to  the  clear  filtrate  as  an  upper 
layer.  If  acetone  is  present,  there  will  be  a  black  zone  (HgS) 
where  the  two  solutions  meet. 

Detection  of  Acetone  in  Urine. — Acidify  200  to  500  cc.  of  urine  with  a  few  drops 
of  sulphuric  acid  and  distil.  Collect  20  to  30  cc.  of  distillate.  This  will  contain 
the  entire  quantity  of  acetone  in  the  urine.  Acetone  thus  obtained  may 
possibly  be  derived  from  aceto-acetic  acid  which  is  often  present  in  human 
urine,  especially  in  a  severe  case  of  diabetes  mellitus.  Distillation  decomposes 
this  acid  into  acetone  and  carbon  dioxide. 

CH3.CO.CH2.CO.OH  =  CH3.CO.CH3  +  CO2. 

Detection  of  Alcohol  and  Acetone  in  Mixtures. — Alcohol  maj^  be  detected  in 
presence  of  acetone  by  Berthelot's  test.  On  the  other  hand,  acetone  may  be 
distinguished  from  alcohol  by  Legal's  or  Penzoldt's  test. 

BITTER  ALMOND  WATER  AND  BENZALDEHYDE 

Bitter  almond  water  (Aqua  Amygdalae  Amar^e  of  the  Phar- 
macopoeia) contains  hydrocyanic  acid.  Only  a  small  portion  of 
this  acid,  however,  is  free  so  that  it  can  be  precipitated  b}'  silver 

nitrate  solution.     The  greater  part  is  combined  as  the  cvano- 

/H 
hydrin  of  benzaldehyde,  CeHs.C^OH,  which  does  not  react  vn.th 

silver  nitrate.    But  potassium  hydroxide  solution  will  decompose 
this  compound. 

CeHsCH  (OH)  CN  +  KOH  =  KCN  +  H2O  +  CeHs.CHO  . 


54  DETECTION   OF   POISONS 

Pure  benzaldehyde,  also  called  hydrocyanic  acid-free  oil  of 
bitter  almonds,  is  not  poisonous.  It  is  oxidized  to  benzoic  acid 
in  the  body  and  eliminated  in  the  urine  partly  as  that  acid  and 
partly  as  hippuric  acid  after  conjugation  with  glycocoll  (amino- 
acetic  acid) : 

(a)   CeHs-CHO      +0  =  CeHs-COOH, 

(/3)    CeHs-COOH  +  H2N-CH2-COOH  =  CeHs-CO-NH-CHj-COOH. 

Benzoic  acid  Glycocoll  furnished  Hippuric  acid 

by  the  organism 

Ordinary  commercial  oil  of  bitter  almonds  contains  hydro- 
cyanic acid  and  is  poisonous  in  proportion  to  the  quantity  of  this 
acid  present. 

Test  for  hydrocyanic  acid  by  shaking  about  2  cc.  of  oil  of 
bitter  almonds  with  20  cc.  of  potassium  hydroxide  solution  and 
making  the  Prussian  blue  test.  When  oil  of  bitter  almonds  is 
mixed  with  other  material,  distil  with  steam  from  a  solution 
acidified  with  tartaric,  or  dilute  sulphuric  acid,  and  test  the  first 
part  of  the  distillate  for  hydrocyanic  acid.  If  benzaldehyde  is 
present,  the  distillate  at  the  same  time  will  be  milky  and  have 
the  characteristic  odor  of  that  compound.  Distil  until  the  drops 
of  water  are  perfectly  clear.  Benzaldehyde  may  be  detected 
with  certainty,  and  at  the  same  time  distinguished  from  nitro- 
benzene which  has  a  somewhat  similar  odor,  by  adding  a  few 
drops  of  potassium  hydroxide  solution  to  the  milky  distillate, 
to  combine  with  any  hydrocyanic  acid,  and  extracting  with 
ether.  The  ether  upon  evaporation  will  deposit  benzaldehyde 
as  globules,  which  can  be  positively  identified  by  conversion  into 
benzoic  acid.  Heat  the  globules  for  a  few  minutes  in  a  small 
flask,  attached  to  a  reflux  condenser,  with  about  10  cc.  of  potas- 
sium dichromate  solution  and  a  little  dilute  sulphuric  acid. 
Cool,  extract  with  ether  and  evaporate  the  ether  solution  in  a 
glass  dish.  When  the  material  contains  benzaldehyde,  this 
residue  will  consist  of  benzoic  acid.  This  substance  may  be 
further  identified  by  its  melting  point  (120-121°),  its  property 
of  subliming  and  the  test  with  ferric  chloride  solution.^ 

^  Dissolve  the  residue  in  a  small  quantity  of  water,  and  neutralize  benzoic  acid 
by  heating  the  solution  to  boiling  with  excess  of  calcium  carbonate.  Filter  and 
add  a  few  drops  of  ferric  chloride  solution.  If  benzoic  acid  is  present,  a  flesh- 
colored  precipitate  of  basic  ferric  benzoate  will  appear.     Tr. 


VOLATILE   POISONS  65 

SYNOPSIS  OF  GROUP  I 
Scherer's  Test  for  Phosphorus  Precedes  Distillation 

The  material  to  be  examined  must  first  be  rendered  uniform 
by  grinding  or  chopping.  Add  sufficient  water  to  thin  the  mass, 
acidify  with  tartaric  acid  and  distil.  If  the  preliminary  test 
for  phosphorus  (Scherer's)  is  positive,  distil  in  the  Mitscherlich 
apparatus;  otherwise  distil  as  usual  with  a  Liebig  condenser. 
It  is  advisable  to  collect  the  distillate  in  two  or  three  fractions. 
Test  the  first  5  to  lo  cc.  of  distillate  for  hydrocyanic  acid, 
chloroform,  ethyl  alcohol,  acetone  and  possibly  also  for  nitro- 
benzene and  iodoform.  Use  the  remainder  of  the  distillate 
in  testing  for  carbolic  acid,  chloral  hydrate  and  carbon 
disulphide. 

Phosphorus. — Phosphorescence  in  Mitscherlich  apparatus 
during  distillation  in  a  dark  room.  Evaporate  distillate  with 
strong  chlorine  water,  or  a  little  fuming  nitric  acid,  and  test  the 
residue  for  phosphoric  acid.  As  an  alternative  procedure, 
examine  the  original  material,  or  at  least  the  Mitscherlich  dis- 
tillate, for  phosphorus  by  the  Blondlot-Dusart  method. 

Hydrocyanic  Acid. — Odor.  Schonbein's  preKminary  test. 
Prussian  blue  test.  Sulphocyanate  test.  Nitroprusside  test. 
Silver  nitrate  test.     Alkaline  phenolphthahn  test. 

CarboUc  Acid. — Odor.  Red  color  with  Millon's  reagent. 
Yellowish  white  precipitate  with  bromine  water.  Violet  color 
with  ferric  chloride  solution. 

Chloroform. — Separation  of  colorless  globules,  when  the 
quantity  is  large.  Odor.  Phenylisocyanide  test,  when  heated 
with  aniline  and  potassium  hydroxide  solution,  Reduces  silver 
nitrate  and  Fehling's  solutions  with  heat.  Red  color  with 
resorcinol  and  potassium  hydroxide  solution.  Blue  color  with 
naphthol  and  potassium  hydroxide  solution. 

Chloral  Hydrate. — Gives  chloroform  reactions.  Brick-red 
precipitate  with  Nessler's  solution  which  in  time  becomes 
yellowish  green.  Gives  chloroform  and  magnesium  formate, 
when  heated  with  magnesium  oxide  and  water.  Test  for 
formate  with  silver  nitrate  or  mercuric  chloride  solution. 


56  DETECTION   OF   POISONS 

Iodoform. — Odor.  Distillate  milky  and  yellowisli  white. 
Ether  extract  of  distillate  leaves  crystals  upon  evaporation, 
gives  chloroform  reactions. 

Nitrobenzene. — Yellowish  globules  with  characteristic  odor. 
Reduced  to  aniline,  when  shaken  with  tin  and  hydrochloric 
acid.     Test  for  aniline. 

Aniline. — Violet  color  with  calcium  hypochlorite  solution. 
Phenyhsocyanide  test,  when  heated  with  chloroform  and 
potassium  hydroxide  solution.  Flesh-colored  precipitate  with 
bromine  water.  Dark  red  color  on  warming  with  Millon's 
reagent. 

Carbon  Bisulphide. — Black  precipitate,  or  only  black  colora- 
tion (PbS),  when  heated  with  lead  acetate  and  potassium 
hydroxide  solutions.  Formation  of  ammonium  sulphocyanate 
by  evaporation  with  concentrated  ammonium  hydroxide  solu- 
tion and  detection  with  ferric  chloride  solution.  Formation 
of  potassium  xanthogenate,  when  shaken  with  alcoholic  solu- 
tion of  potassium  hydroxide  and  detection  with  copper  sulphate 
solution. 

Ethyl  Alcohol. — Iodoform  test.  Odor  of  ethyl  benzoate, 
when  shaken  with  benzoyl  chloride  and  sodium  hydroxide  solu- 
tion. Green  color  and  aldehyde  odor,  when  heated  with  potas- 
sium dichromate  and  hydrochloric  acid.     Vitali's  test. 

Acetone. — Gives  iodoform,  even  in  the  cold,  with  iodine 
and  potassium  hydroxide  or  ammonium  hydroxide  solution. 
LegaFs'^est.     Indigotin  test.     Reynold's  test. 


CHAPTER  II 

NON-VOLATILE  POISONS  ^ 

Alkaloids,  Glucosides  and  Synthetic  Compounds  Non-volatile  from  Acid 
Solution  with  Steam 

Put  a  portion  of  finely  chopped  material  into  a  large  flask, 
and  thoroughly  mix  with  two  or  three  times  as  much  absolute 
alcohol. 2  Add  enough  tartaric  acid  solution  to  give  the  mixture 
a  distinct  acid  reaction  after  shaking.  Laboratory  experiments 
usually  require  20  to  30  drops  of  10  per  cent,  tartaric  acid  solu- 
tion. Avoid  a  large  excess  of  tartaric  acid,  since  it  may  act 
as  an  objectionable  impurity  in  the  ether  extract,  owing  to  its 
solubihty  in  that  solvent.  Connect  the  flask  with  a  glass  tube 
(80  to  100  cm.  long)  serving  as  a  reflux  cooler.  Frequently 
sha,ke  and  heat  10  to  15  minutes  upon  the  water-bath.  In  the 
extraction  of  a  large  quantity  of  material  from  a  cadaver,  con- 
nect the  flask  with  an  upright  Liebig  condenser  used  as  a  reflux 
cooler  (Fig.  10).  Cool  the  flask  contents  and  filter  to  remove 
fat  and  other  insoluble  matter.  Wash  the  residue  with  alcohol. 
Evaporate  the  filtrate,  which  must  have  an  acid  reaction,  to  a 
thin  syrup  in  a  glass  dish  upon  the  water-bath.  Thoroughly 
mix  this  residue  with  100  cc.  of  cold  water.  Usually  this 
causes  an  abundant  separation  of  fat  and  resinous  matter, 
especially  when  parts  of  a  cadaver  are  examined.  Filter  and 
evaporate  the  filtrate  to  dryness,  or  to  a  syrup,  upon  the  water- 

^  The  isolation  of  these  toxic  substances  from  cadaveric  material,  food,  etc., 
is  necessary  before  tests  establishing  their  presence  can  be  made,  ^fixtures  used 
for  laboratory  practice,  consisting  of  dog  biscuit,  meat,  comminuted  organs  (liver, 
kidneys,  spleen),  sausage  meat,  etc.,  with  any  of  the  poisons  of  this  group,  should 
be  examined  according  to  the  method  outlined  above. 

^  Commercial  alcohol  usually  contains  basic  compounds,  the  presence  of  which 
is  objectionable.  They  should  be  removed  by  adding  tartaric  acid  to  the  alcohol 
and  distiUing.  Alcohol  should  not  be  used  in  toxicological  analysis,  unless  an 
actual  test  has  shown  it  to  be  free  from  such  impurities.     Tr. 

57 


58 


DETECTION   OF   POISONS 


bath.  ,  Thoroughly  mix  this  residue  with  absolute  alcohol.  As 
a  result  of  this  treatment,  a  whitish  substance,  which  is  more  or 
less  viscous  or  slimy,  usually  remains  undissolved.  This  resi- 
due, which  consists  chiefly  of  protein  substances  (albumin, 
albumoses  and  peptones),  dextrin-like  compounds  and  in  part 
also  of  inorganic  salts,  frequently  be- 
comes granular  upon  standing.  Tartrates 
of  the  alkaloids  and  other  organic  poisons 
are  dissolved.  The  larger  the  quantity 
of  absolute  alcohol  used,  the  more  com- 
plete the  precipitation  of  those  substances 
which  interfere  more  or  less  with  the  detec- 
tion of  organic  poisons.  Again  evaporate 
the  filtered  alcoholic  solution  upon  the 
water-bath,  and  dissolve  the  residue  in 
about  50  cc.  of  water.  If  the  solution  is 
not  perfectly  clear,  filter  through  a  moist- 
ened paper. 

The  result  of  this  procedure  is  a  solution 
containing  alkaloidal  tartrates  and  other 
organic  substances  belonging  to  this  group. 
This  solution  should  have  an  acid  reac- 
tion and  be  practically  free  from  protein 
substances,  fat,  resinous  bodies  and  color- 
ing matter.  If  the  solution  fulfils  these 
requirements,  it  is  ready  to  be  examined 
for  organic  poisons  according  to  the  "Stas- 
Otto  "  method.  The  utmost  care  must  be 
taken  in  preparing  this  solution,  because 
definite  conclusions  cannot  be  drawn  from 
the  uncertain  tests  given  by  impure  material. 

When  the  material  is  a  powder  mixed  with  cane-  or  milk- 
sugar,  it  is  usually  possible,  after  the  aqueous  solution  has  been 
acidified  with  tartaric  acid,  to  extract  directly  with  ether  and 
continue  according  to  the  Stas-Otto  method. 

Frequently  in  suspected  poisoning  an  examination  of  beer, 
wine,  black  coffee,  infusion  of  tea,  food,  etc.,  is  necessary.     In 


Fig.    10. — ^Liebig    Con- 
denser as  Reflux  Cooler. 


NON-VOLATILE   POISONS 


J9 


such  cases  the  process  outlined  above  may  often  be  considerably 
shortened.  Acidify  the  material  with  aqueous  tartaric  acid 
solution,  if  necessary,  and  evaporate  in  a  glass  dish  upon  the 
water-bath.  Treat  the  residue  thoroughly  with  absolute  alcohol 
and  fdter.  Evaporate  the  filtrate  upon  the  water-bath  and 
dissolve  the  residue  in  tepid  water.  Filter  this  solution,  if 
necessary,  and  then  examine  according  to  the  Stas-Otto  process. 


Fig.  II. — Separating  Funnels  and  Glass  Crystallizing  Dishes. 

STAS-OTTO  PROCESS 

A.  Examination  of  Ether  Extract  of  Tartaric  Acid  Solution 

Thoroughly  extract  the  acid  aqueous  solution  (see  process  of 
preparation  described  above)  two  or  three  times  wdth  ether, 
using  each  time  about  the  same  quantity  of  solvent.     Employ 


60  DETECTION   OF  POISONS 

a  separating  funnel  for  this  purpose  (Fig.  ii).  Pour  the  com- 
bined ether  extracts  into  a  dry  flask  loosely  stoppered.  If  the 
solution  stands  for  i  or  2  hours  at  rest,  a  few  drops  of  water 
usually  settle  out.  Decant  the  ether  solution  and  pour  through 
a  dry  filter.  Slowly  evaporate  this  solution  in  a  small  glass 
dish  upon  a  water-bath  previously  heated  slightly  above  35°. 
Do  not  have  gas  burning  during  this  operation!  Examine  the 
residue  as  described  below.  An  excellent  method  of  evaporat- 
ing ether  consists  in  setting  a  small  glass  dish  (8  to  10  cm.  in 
diameter)  upon  a  hot  water-bath  and  dropping  the  filtered  ether 
extract  into  it  as  fast  as  the  solvent  evaporates.  Thus  a  large 
quantity  of  extract  may  be  evaporated  in  a  small  dish.  The 
advantage  of  this  method  is  the  ease  with  which  the  residue  can 
be  removed  for  the  various  tests.  The  residue  is  usually  quite 
small  and  it  is  not  advisable  to  have  it  distributed  over  too 
large  a  surface. 

Examine  the  residue  from  the  ether  extract  for  the  following 
substances : 

Picrotoxin  Caffeine  Antipyrine 

Colchicin  Acetanilide  Salicylic  Acid 

Picric  Acid  Phenacetine  Veronal 

Evaporation  of  the  ether  extract,  even  in  the  absence  of 
members  of  the  group,  usually  leaves  a  more  or  less  viscous 
residue,  containing  tartaric  and  lactic  acids  as  well  as  fatty, 
resinous  and  colored  substances.  This  is  especially  so  in  analy- 
ses of  cadaveric  material.  Moreover  ether  extracts  from  aque- 
ous solutions  certain  metallic  salts,  for  example,  mercuric 
cyanide^  and  chloride. 

^  Ether  will  extract  mercuric  cyanide  to  some  extent  from  a  tartaric  acid  solu- 
tion which  is  not  too  dilute.  For  instance,  it  wiU  remove  appreciable  quantities 
from  100  cc.  of  o.i  per  cent,  mercuric  cyanide  solution,  but  the  extraction  will  not 
be  complete.  The  solution  after  five  extractions  will  still  give  a  distinct  test  for 
mercury.  Ether  wiU  not  remove  even  a  trace  of  mercuric  cyanide  from  o.oi  per 
cent,  solution.  To  test  for  cyanide,  add  ammonium  sulphide  solution  to  the 
ether  residue.  This  wiU  precipitate  mercuric  sulphide  and  the  filtrate  will  con- 
tain ammonium  sulphocyanate  (see  hydrocyanic  acid,  page  22). 


NON-VOLATILE   POISONS  01 

First,  note  the  general  appearance  and  taste  of  the  residue. 
Then  examine  it  with  a  microscope.  Very  defmite  conclusions 
as  to  the  presence  or  absence  of  certain  substances  can  frequently 
be  drawn.  A  very  bitter  residue  should  be  examined  carefully 
for  picrotoxin  and  colchicin.  If  there  is  a  pronounced  yellow 
color,  the  examination  should  include  picric  acid  also.  Veronal 
is  colorless  and  has  a  very  bitter  taste.  A  tasteless,  or  only 
faintly  bitter,  residue  probably  does  not  contain  these  sub- 
stances and  should  be  examined  for  acetaniUde,  antipyrine, 
caffeine,  phenacetine  and  salicylic  acid. 

The  residue  from  evaporation  of  the  ether  extract  may  con- 
tain the  following  substances: 

Picrotoxin. — Usually  a  thick  syrup  which  gradually  solidifies 
and  becomes  crystalline.     Tastes  intensely  bitter. 

Colchicin. — Yellowish,  amorphous  residue  which  does  not 
become  crystalline.  Tastes  intensely  bitter.  Dissolves  in 
water  with  a  yellowish  color,  which  increases  in  intensity  on 
addition  of  a  few  drops  of  dilute  hydrochloric  acid. 

Picric  Acid. — Usually  appears  as  a  syrup  which  gradually 
solidifies  and  becomes  crystalHne.  Tastes  very  bitter.  Resi- 
due intensely  yellow,  giving  yellow  aqueous  solutions  not  in- 
tensified by  hydrochloric  or  sulphuric  acid. 

AcetaniUde.- — ^Leaflets  or  flattened  needles.  Has  a  faint, 
burning  taste  bu-t  is  not  bitter. 

Phenacetine. — Inodorous  and  tasteless  leaflets  and  small 
needles. 

Antipyrine. — Residue  a  syrup  which  is  rarely  crystaUine. 
Tastes  mildly  bitter.     Very  easily  soluble  in  water. 

Caffeine. — Residue  composed  of  shining  needles  frequently 
in  radiating  clusters.     Tastes  mildly  bitter. 

Salicylic  Acid. — CrystalHzes  frequently  in  long  needles. 
Tastes  harsh  and  at  the  same  time  sweet  and  acid. 

Veronal. — Crystalline  needles  ha\dng  an  agreeable,  bitter 
taste. 

PICROTOXIN 

Picrotoxin,  C30H34O13,  the  poisonous  principle  of  Cocculus  indicus,  the  fruit  of 
Menispermum  Cocculus,  crystallizes  from  hot  water  in  long  colorless  needles 


62  DETECTION   OF  POISONS 

melting  at  199-200°.  It  dissolves  with  difiSculty  in  cold  water  but  more  readily  in 
hot  water  or  alcohol.  It  is  sHghtly  soluble  in  ether  but  freely  soluble  in  chloro- 
form, amyl  alcohol  and  glacial  acetic  acid.  Its  alcoholic  solution  is  neutral  and 
laevo-rotatory.  Picrotoxin  has  a  very  bitter  taste.  It  is  not  as  readily  soluble  in 
acids  as  in  pure  water,  but  is  soluble  in  caustic  alkalies  and  aqueous  ammonia, 
forming  unstable,  salt-like  compounds  which  do  not  crystallize.  Picrotoxin 
behaves  toward  strong  bases  as  if  it  were  a  weak  acid.  Heated  to  boiling  with 
twenty  times  its  volume  of  benzene,  it  is  decomposed  into  picrotoxinin  and  picro- 
tin.     The  former  passes  into  solution  but  picrotin  is  almost  completely  insoluble : 

C30II34O13  =  CiBHieOe  +  CibHisOv 

Picrotoxin       Picrotoxinin       Picrotin 

Chloroform  brings  about  this  cleavage  even  more  easily.  On  the  other  hand, 
if  picrotoxinin  and  picrotin  in  molecular  proportions  are  dissolved  in  hot  water, 
picrotoxin  crystallizes  out  as  the  solution  cools.  Treated  with  bromine  direct  or 
dissolved  in  water  or  ether,  picrotoxin  is  first  split  into  picrotoxinin  and  picrotin. 
The  former  is  immediately  converted  into  monobromo-picrotoxinin,  CiBHiBBrOe, 
but  picrotin  remains  almost  unchanged.  Monobromo-picrotoxinin  is  soluble 
with  difi&culty  in  water  but  is  reduced  by  zinc  dust  and  acetic  acid  to  picrotoxinin. 
Picrotin  is  almost  non-toxic,  whereas  picrotoxinin  has  a  very  poisonous  action. 
Picrotoxin  is  a  powerful  convulsive  poison,  standing  in  its  action  between  cicu- 
toxin  and  strychnine. 

R.  Meyer  and  P.  Bruger^  regard  picrotoxin  as  a  complex  of  the  two  compounds, 
picrotin  and  picrotoxinin,  crystallizing  together  in  definite  but  not  molecular 
proportion,  and  not  as  a  molecularly  constituted  chemical  compound. 

Detection  of  Picrotoxin 

1.  Fehling's  Test. — Dissolve  picrotoxin  in  a  small  test-tube, 
using  10-20  drops  of  very  dilute  sodium  hydroxide  solution. 
Add  a  few  drops  of  Fehling's  solution^  and  warm  but  do  not 
shake.  A  red  or  yellowish  red  precipitate  forms  and  settles  to 
the  bottom.  If  the  ether  residue,  not  too  Httle  of  which  should 
be  taken,  fails  to  give  a  clear  solution  in  very  dilute  sodium 
hydroxide  solution,  filter  through  moistened  paper  and  examine 
the  filtrate  with  Fehling's  solution. 

2.  Ammoniacal  Silver  Test.- — ^Warm  picrotoxin  with  aqueous 
silver  nitrate  solution  containing  a  slight  excess  of  ammonium 
hydroxide  solution.  The  reducing  action  of  picrotoxin  will 
produce  a  black  precipitate  of  metallic  silver,  or  a  dark  brown 
color  when  only  traces  are  present. 


^  Berichte  der  Deutsehen  chemischen  Gesellschaft  31,  2958  (i{ 
^  Fehling's  solution  heated  by  itself  should  not  give  a  precipitate  of  cuprous 
oxide. 


NON-VOLATILE   POISONS  63 

3.  Oxidation  Test. — Picrotoxin,  treated  with  a  little  con- 
centrated sulphuric  acid  in  a  porcelain  dish,  first  becomes 
orange-red  and  then  dissolves  when  stirred  forming  a  reddish 
yellow  solution.  A  drop  of  potassium  dichromate  solution  will 
produce  a  red-brown  color  around  the  margin  of  the  drop.  If 
the  two  Kquids  are  thoroughly  mixed,  there  is  an  immediate 
dirty  brown  color  which  passes  into  green  on  long  standing. 

A  green  color  alone  is  without  significance,  since  many  organic  substances 
capable  of  reducing  chromic  acid  to  chromic  oxide  produce  the  same  result. 

4.  H.  Melzer's^  Test. — Put  some  picrotoxin  upon  a  watch 
glass  and  add  i  or  2  drops  of  a  mixture  of  benzaldehyde  and 
absolute  alcohol.  Careful  addition  of  a  drop  of  concentrated 
sulphuric  acid  will  produce  a  distinct  red  color.  If  the  watch 
glass  is  tilted,  red  streaks  will  run  from  the  substance  through 
the  liquid. 

Use  a  freshly  prepared,  20  per  cent,  solution  of  benzaldehyde  in  absolute  alco- 
hol. Benzaldehyde  alone  gives  a  yellowish  brown  color  with  concentrated 
sulphuric  acid.  Alcohol  is  added  as  a  diluent  to  diminish  this  color  as  much  as 
possible.  Under  these  conditions  the  solution  has  a  light  yellow  color,  and  the 
dark  red  tint  caused  by  picrotoxin  is  very  clearly  defined.  This  red  color  is 
unstable  and,  beginning  at  the  margin,  gradually  fades  into  a  pale  pink  or  \aolet. 
H.  Kreis-  has  found  that  cholesterine  and  phytosterine'  give  similar  colors  with 
Melzer's  reagent. 

5.  Langley's  Test. — Mix  picrotoxin  with  about  3  times 
the  quantity  of  potassium  m'trate,  and  moisten  the  mixture 
with  the  smallest  possible  quantity  of  concentrated  sulphuric 
acid.  Then  add  strong  sodium  hydroxide  solution  in  excess 
and  an  intense  red  color  will  appear. 

Detection  of  Picrotoxin  in  Beer 

First,  neutralize  the  beer  mth  magnesium  oxide.  Then  evaporate  500  cc.  or 
more  to  a  syrup  upon  the  water-bath.  Digest  this  residue  vdih  4  or  5  times  its 
volume  of  alcohol  and  evaporate  the  alcohoHc  extract.  Dissolve  the  residue  in 
hot  water  and  filter  the  solution  through  a  moistened  paper.     Acidify  the  filtrate 

1  Zeitschrift  fiir  analytische  Chemie  37,  351  and  747  (1898). 

2  Chemiker-Zeitung  33,  21  (1899). 

^  A  substance  very  similar  to  cholesterine,  and  named  paracholesterine  or 
phytosterine,  is  found  in  the  seeds  of  certain  plants.  (Perkin  and  Kipping, 
Organic  Chemistry,  page  608.) 


64  DETECTION   OF   POISONS 

with  dilute  sulphuric  acid  and  extract  repeatedly  with  ether,  or  better  with  chlo- 
roform.    Evaporate  these  extracts  and  test  the  residue  for  picrotoxin. 

Should  the  residue  from  the  ether  or  chloroform  be  too  impure,  dissolve  it  again 
in  hot  water,  filter,  evaporate  and  extract  with  ether  or  chloroform.  To  purify 
picrotoxin  further,  precipitate  colored  substances  from  its  aqueous  solution  with 
lead  acetate,  filter  and  remove  lead  from  the  filtrate  by  hydrogen  sulphide.  The 
filtrate  from  lead  sulphide  upon  evaporation,  or  extraction  with  ether  or  chloro- 
form, will  give  nearly  pure  picrotoxin.  The  very  bitter  taste  of  picrotoxin  as 
weU  as  its  strong  tendency  to  crystalUze  are  additional  characteristics  of  this 
substance. 

COLCHICm 

Colchicin,  C22H26NO6,  an  alkaloid  occurring  in  all  parts  of  the  meadow  saffron, 
Colchicum  autumnale,  is  a  yellowish,  amorphous  powder  which  is  poisonous  and 
very  bitter  to  the  taste.  It  is  freely  soluble  in  water,  alcohol,  and  chloroform,  less 
so  in  either,  and  benzene,  and  almost  insoluble  in  petroleum  ether.  Solutions  of 
colchicin  have  a  more  or  less  yellowish  color  which  becomes  more  pronounced 
upon  addition  of  acids  or  alkalies.  These  solutions  have  very  faint  basic  proper- 
ties. Consequently  ether  or  chloroform,  but  not  benzene  nor  petroleum  ether,  will 
extract  colchicin  from  an  acid,  aqueous  solution.  Upon  evaporation  of  the  sol- 
vent, colchicin  will  appear  as  a  yellowish,  sticky  residue  resembUng  a  resin  or 
varnish.  Heated  with  water  containing  sulphuric  acid,  colchicin  splits  into  col- 
chicein  and  methyl  alcohol.  BoiHng  the  alkaloid  1.5-2  hours  with  60  parts  of  i 
per  cent,  hydrochloric  acid  will  produce  the  same  result: 

C22H26NO6  +  H2O  =  C21H23NO6  +  CH3.OH 
Colchicin  Colchicein    Methyl  Alcohol 

On  the  other  hand,  colchicin  is  formed  when  colchicein  is  heated  to  100°  with 
sodium  methylate  (CHs.ONa)  and  methyl  iodide  (CH3.I).  Since  colchicein  on 
treatment  with  hydriodic  acid  yields  three  molecules  of  methyl  iodide,  colchicein 
as  well  as  colchicin  contains  three  methoxyl  groups.  Heated  with  strong  hydro- 
chloric acid,  colchicein  loses  acetic  acid  and  passes  into  trimethyl-colchicinic 
acid.  Consequently  colchicein  and  colchicin  contain  an  acetyl  group  (CH3.CO  — ). 
The  formula  of  colchicin,  that  is  to  say,  of  methyl-colchicein,  may  be  written  as 
follows : 

CH30\  /NH.CO.CH3 

CH3O— Cl6H9< 

CH3O/  \CO.OCH3 

Detection  of  Colchicin 

Aqueous  colchicin  solutions,  especially  in  presence  of  dilute 
mineral  acids,  have  a  yellow  color.  Unless  the  ether  residue  has 
this  characteristic,  colchicin  is  absent. 

I.  Tannic  Acid  Test. — This  reagent  will  precipitate  colchicin 
from  aqueous  solution,  if  not  too  dilute,  as  white  flocks.  This 
test,  however,  is  not  characteristic  of  colchicin. 


NON-VOLATILE    POISONS  05 

2.  Nitric  Acid  Test. — Nitric  acid  (Sp.  gr.  1.4  =  66  per  cent.) 
dissolves  colchicin  with  a  dirty  violet  color  which  soon  changes, 
when  stirred,  to  brownish  red  and  finally  to  yellow.  Addition  of 
dilute  sodium  or  potassium  hydroxide  solution,  until  the  reaction 
is  alkaline,  produces  an  orange-yellow  or  orange-red  color. 

3.  Sulphuric  Acid  Test.— Concentrated  sulphuric  acid  dis- 
solves colchicin  with  an  intense  yellow  color.  A  drop  of  nitric 
acid  added  to  such  a  solution  produces  a  green,  blue,  violet  and 
finally  a  pale  yellow  tone.  Excess  of  potassium  hydroxide  solu- 
tion will  now  bring  out  an  orange-red  color.  Erdmann's 
reagent  (see  page  313)  dissolves  colchicin  with  a  blue  to  violet 
color, 

4.  Hydrochloric  Acid  Test. — Concentrated  hydrochloric  acid 
dissolves  colchicin  with  an  intense  yellow  color.  Add  two  drops 
of  ferric  chloride  solution  and  heat  the  mixture  2-3  minutes  in  a 
test-tube.  The  color  deepens  and  the  solution  on  cooling, 
especially  if  diluted  with  the  same  volume  of  water,  becomes 
green  or  olive-green.  Finally  shake  the  solution  with  a  few 
drops  of  chloroform.  This  solvent  becomes  yellowish  brown,  or 
garnet-red,  and  the  aqueous  solution  retains  its  green  color. 
Zeisel's  reaction. 

Purification  of  the  Residue  Containing  Colchicin 

To  isolate  as  pure  colchicin  as  possible  from  the  yellow 
residue,  extract  with  warm  water.  Filter  the  solution  and, 
when  cold,  extract  it  first  with  petroleum  ether.  This  will 
remove  fatty,  resinous  and  colored  impurities  but  not  colchicin. 
Then  extract  with  chloroform.  Or  precipitate  colchicin  from 
aqueous  solution,  which  must  not  be  too  dilute,  with  tannic 
acid.  Collect  this  precipitate  upon  a  filter  and  wash  with  cold 
water.  Mix  the  moist  precipitate  with  freshly  precipitated, 
washed  lead  hydroxide.  Dry  the  mixture,  grind  to  a  powder 
and  extract  with  chloroform.  Evaporation  of  the  solvent  will 
leave  nearly  pure  colchicin. 

PICRIC  Acm 

Picric  acid,  or  2,4,6-trinitrophenol,   crj^stallizes  from  water  in  light  yellow 
leaflets  and  from  ether  in  lemon-jeUow,  rhombic  prisms.     It  melts  at  122.5°. 
5 


66  DETECTION   OF   POISONS 

Though  soluble  in  cold  water  with  difficulty,  picric  acid  dissolves  freely  in  hot 

water,  as  weU  as  in  alcohol,  ether  and  benzene.     Aqueous  solutions  have  an  acid 

PjTT  reaction,  a  very  bitter  taste  and  dye  animal  fibers  fast  yellow. 

I  Material  containing  picric  acid  has  a  yeUow  or  yellowish 

C  green  color. 

'^\  Physiological   Action    and  Elimination. — Picric  acid  is 

"^         I        II  ^  quite  an  active  poison.     Taken  internally  it  produces  a 

HC      CH  striking  yellow  pigmentation  first  of  the  conjunctiva  and 

\/  then  of  the  entire  skin,  usually  designated  as  "picric  acid 

C  icterus."     Picric  acid  and  its  salts  like  most  nitro-com- 

Jj-p.  pounds  decompose  the  red  blood-corpuscles  forming  met- 

hsemoglobin.      Consequently  it  is  a  blood-poison.     At  the 

same  time  it  irritates  the  central  nervous  system  and  causes  convulsions. 

Finally  it  exercises  its  power  of  precipitating  proteins  in  acid  solution.     This  is 

especially  noticeable  in  those  organs  of  the  body,  for  example,  the  stomach  and 

OH  kidneys,  which,  owing  to  necrotic  tissue  changes,  have  an 

I  acid  or  only  a  faintly  alkaline  reaction.     The  organism  re- 

^  duces  picric  to  picraminic  acid  which  does  not  so  readily 

Q  Tyr Q       p j^jj  precipitate   protein.     By   thus   changing   picric   acid   the 

I        II  organism  rids  itself  of  the  poison.     In  picric  acid  poisoning 

HC       CH         the  urine  has  a  marked  red  color  owing  to  formation  of 

^^  picraminic  acid.     Some  picric  acid  passes  into  the  urine 

r  unchanged.     Elimination   is   slow.     In   one   case    (see   R. 

^"02  Kobert,  "Intoxikationen"),  after  administration  of  a  single 

Picraminic  acid      dose  of  I  gram  of  picric  acid,  its  presence  in  the  urine  could 

be  recognized  for  6  days.      The  urine  was  ruby  red,  clear,  acid  and  free  from 

albumin  and  bUe-constituents.     Picric  acid  was  also  easily  detected  in  the  fasces. 

Detection  of  Picric  Acid 

Material  containing  picric  acid  has  a  more  or  less  yellow  or 
yellowish  green  color.  Aqueous,  alcoholic  and  ethereal  solu- 
tions show  the  same  color.  Finely  divided  animal  material 
should  be  extracted  several  hours  under  a  return-condenser  with 
alcohol  containing  hydrochloric  acid  to  decompose  compounds  of 
picric  acid  with  albumins  and  thus  bring  the  acid  into  solution. 
Filter  and  evaporate  such  an  alcoholic  extract  upon  the  water- 
bath.  Treat  the  residue,  which  is  yellow,  yellowish  green,  or 
frequently  yellowish  red  or  reddish  brown,  with  warm  water  and 
filter  the  extract.  The  filtrate  itself  may  be  tested  directly 
for  picric  acid,  or  it  may  first  be  extracted  as  usual  with  con- 
siderable ether.  The  following  tests  may  then  be  applied  to  the 
residue  left  on  evaporating  the  ether  extract: 


NON-VOLATILE   POISONS  67 

1.  Isopurpuric  Acid  Test. — Gently  heat  (50-60°)  an  aqueous 
solution  of  picric  acid  with  a  few  drops  of  saturated,  aqueous 
potassium  cyanide  solution  (i  :  2) .  The  solution  will  become  red 
owing  to  formation  of  potassium  isopurpurate.  One  milligram 
of  picric  acid,  dissolved  in  5  cc.  of  water,  will  give  a  distinct  test. 

Isopurpuric  acid  does  not  exist  in  the  free  state  but  is  present  in  this  test  as 
the  potassium  salt.     Nietzki  and  Petri^  regard  isopurpuric  acid   (CsHjOeNs) 
as    a    dicyano-picraminic    acid  =  5-oxy-6-amino-2,4-dinitro-isophthalic  nitrile; 
whereas  Borsche'^  considers  it  a  dicyano-dinitro-oxy-;3-phenyl  hydroxylamine: 
OH  OH 

C  C 

/\  /\ 

O2N— C     C— NH2  O2N— C      C— NH.OH 

I       II  I        II 

NC— C     C— CN  NC— C     C— CN 

\/  \/ 

c  c 

I  I 

.  N02  N02 

Nietzki-Petri  Borsche 

2.  Picraminic  Acid  Test. — (a)  Heat  picric  acid  solution  with 
a  few  drops  of  sodium  hydroxide  solution  and  glucose.  Picra- 
minic acid,  formed  by  reduction  of  picric  acid,  colors  the  solu- 
tion deep  red.  Avoid  excess  of  sodium  hydroxide  solution, 
otherwise  there  will  be  a  red  color  due  solely  to  the  action  of  the 
alkali  upon  glucose. 

(i(3)  The  test  may  also  be  made  by  warming  picric  acid 
solution  with  a  few  drops  of  sodium  hydroxide  and  ammonium 
sulphide  solutions.  This  will  reduce  picric  acid  and  produce  a 
red  color. 

In  both  reactions  (a  and  0)  picric  acid  is  reduced  to  picraminic 
acid,  2-amino-4,6-dinitro-phenol : 

OH  OH 

i  i 

/\  /\ 

O2N— Ce      2C— NO2  O2N— Ce       2C— NHo 

I  II  +  6H  =  I  II  "  +  2H2O. 

HC        CH  HC        CH 

c  c 

I  I 

N02  N02 

Picric  acid  Picraminic  acid 

^  Berichte  der  Deutschen  chemischen  Gesellschaft  SS)  1788  (1900). 
^  Ibid.,  ss,  2718  and  2995  (1900). 


68  DETECTION   OF   POISONS 

The  presence  of  fat  and  other  impurities  materially  influences 
this  test. 

3.  Dyeing  Test. — Dissolve  the  substance  containing  picric 
acid  in  hot  water  and  put  white  threads  of  wool,  silk  and  cotton 
in  the  solution.  In  a  few  hours  (12  to  24)  remove  the  threads 
and  thoroughly  rinse  in  pure  water.  If  picric  acid  is  present, 
the  wool  and  silk  will  be  dyed  yellow  but  not  the  cotton.  In 
other  words,  picric  acid  is  not  fast  upon  vegetable  fibers  like 
cotton.  Picric  acid,  diluted  i :  100,000,  will  still  produce  a  yel- 
low color  upon  wool. 

4.  Ammoniacal  Copper  Test. — ^Add  a  few  drops  of  ammonia- 
cal  copper  sulphate  solution  (copper  sulphate  solution  and  an 
excess  of  ammonia)  to  an  aqueous  picric  acid  solution,  A  yel- 
lowish green  precipitate,  consisting  of  hexagonal  needles  with  a 
polarizing  action  upon  light,  will  appear.  Picric  acid,  diluted 
1 :  80,000,  will  give  this  test. 

ACETANILIDE 

•j^TT pQ pjj  Acetanilide  crystallizes  in  colorless  and  inodorous,  shin- 

I  ing  leaflets.     It  has  a  faint,  burning  taste;  melts  at  113  to 

C  114°;  is  soluble  in  230  parts  of  cold  water,  in  about  22  parts 

^\  of  boiling  water  and  in  3.5  parts  of  alcohol;  and  is  freely 

I        II  soluble  in   ether   and   still  more  so  in  chloroform.      All 

HC      CH  acetanilide  solutions  are  neutral.      Heated  to  boiling  with 

\./  potassium   hydroxide  solution  (I)   and  also  with  fuming 

^  hydrochloric  acid  (II),  acetaniUde  is  decomposed  into  its 

constituents : 

I.  CeHs.NH.CO.CHs  +  KOH  =  CeHs.NHz  +  CH3.CO.OK. 

II.  CeHs.NH.CO.CHa  +  HCl  +  H2O  =  CeHj.NHa.HCl  +  CH3.COOH. 

Physiological  Action. — Being  an  aniline  derivative,  acetanihde  has  the 
poisonous  properties  of  thai  amine  though  in  less  degree.  R.  Robert  ("in- 
toxikationen")  refers  to  several  instances  of  acetanilide  poisoning  which  did 
not  terminate  fatally.  In  one  case  a  student  took  a  teaspoonful  of  the  drug. 
There  was  stupor,  uneasiness,  marked  cyanosis  and  lowering  of  the  pulse. 
A  purgative  and  restorative  (stimulant)  were  used  but  there  was  considerable 
exhaustion  for  several  days.  The  picture  was  nearly  the  same  in  the  case  of 
a  man  who  took  2  grams  of  antifebrine  daily  for  2  days  in  succession. 

Preparation. — Boil  aniline  and  glacial  acetic  acid  together  for  several  hours 
under  a  return-condenser: 

CeHB-NHz  -t-  CH3-COOH  =  CeHfi-NH-CO-CHa  +  H2O. 


NON-VOLATILE   POISONS  69 

Detection  of  Acetanilide 

Ether  or  chloroform  will  extract  acetanilide  completely  from 
an  acid  aqueous  solution. 

1.  Indophenol  Test. — Boil  acetanilide  with  about  4  cc.  of 
fuming  hydrochloric  acid  and  evaporate  to  a  few  drops  (about 
10).  Cool  and  add  4  cc.  of  saturated,  aqueous  carbolic  acid 
solution.  A  few  drops  of  calcium  hypochlorite  solution  will 
produce  a  violet-red  color.  In  time  the  color  will  become  deeper, 
especially  if  the  mixture  is  shaken.  Then  carefully  add  ammon- 
ium hydroxide  solution  as  a  surface-layer  which  will  take  on  a 
permanent  indigo-blue  color. 

The  indigo-blue  color  is  characteristic  of  acetanilide  only 
when  preceded  by  the  red-violet  color,  since  a  mixture  of  aque- 
ous phenol  and  hypochlorite  solution  gives  a  blue  color  with 
ammonia  (see  carbolic  acid). 

Phenacetine  also  gives  the  indophenol  test. 

2.  Phenylisocyanide  Test. — Boil  acetanilide  with  5-6  cc. 
of  alcoholic  potassium  hydroxide  solution.  Cool,  add  2  or  3 
drops  of  choloroform  and  again  heat.  The  offensive  odor  of 
phenylisocyanide  will  be  developed. 

Potassium  hydroxide  decomposes  acetanilide  into  aniline 
and  potassium  acetate  (see  Reaction  I  above).  The  former 
with  chloroform  gives  phenylisocyanide. 

3.  Calcium  Hypochlorite  Test. — Boil  acetanilide  a  few  min- 
utes with  alcoholic  potassium  hydroxide  solution  as  in  test  2. 
Dilute  with  water  and  extract  aniline  with  ether.  This  sol- 
vent upon  evaporation  will  deposit  aniline  as  an  oily  liquid. 
Dissolve  the  latter  in  water  and  test  with  calcium  hypochlorite. 

Examination  of  Acetanilide  Urine ^ 

Scarcely  more  than  traces  of  unaltered  acetanilide  appear  in  urine  even  after 
large  doses.  The  most  essential  change  occurring  in  the  body  is  oxidation  of  the 
benzene  ring  which  produces  aceto-para-aminophenol.  This  Uke  most  phenols 
forms  a  conjugate  sulphuric  acid  and  appears  in  the  urine  as  a  salt  of  aceto-para- 
aminophenyl  sulphuric  acid : 

^  To  stud}''  the  behavior  of  acetanilide  in  the  bod}',  take  at  night  0.3  gram  of 
this  substance  at  a  dose  twice  in  the  course  of  3  hours  and  examine  the  urine 
passed  in  the  next  12  hours. 


70  DETECTION   OF  POISONS 

H  OH  O— SO2— OH 

I  -^  I  I 

c  c  c 

/\  /\  /-x 

HC      CH  HC      CH  Txp-  HC      CH 

I       II  +0=       I        II  4."°\so,-       I        II 

HC      CH  oxidation  HC      CH  ^  Tin/         ~  HC      CH 

\/  \/  ^^  \/ 

C  C  conjugation  C 

I  I  I 

NH.CO.CH3  NH.CO.CH3  NH.CO.CH3 

Acetanilide  Aceto-p-aminophenol  Aceto-p-aminophenyl  sulphuric  acid 

To  some  extent  also,  a  conjugate  glycuronic  acid  of  aceto-para-aminophenol  is 

formed.     These  coinpounds,  heated  with  concentrated  hydrochloric  acid,  give 

para-aminophenol  which  can  be  detected  by  the  indophenol  test  previously 

described. 

O.SO2.OH  OH 

I  I 

C  C 

/\  /\ 

HC     CH  HC     CH 

I  II  +  2H2O  =  H2SO4  +  CH3.COOH  +  I  II 
HC     CH  HC      CH 

\/  \/ 

c  c 

I  I 

NH.C0.CH3  NH2 

p-aminophenol 

Such  urine,  boiled  a  few  minutes  with  concentrated  hydrochloric  acid,  will 
usually  give  the  indophenol  test.  But  the  test  will  be  more  certain,  if  para- 
aminophenol  is  first  isolated.  Boil  a  larger  quantity  of  urine  (300  to  500  cc.)  a 
few  minutes  with  about  10  cc.  of  concentrated  hydrochloric  acid.  Then  add  an 
excess  of  sodium  carbonate  and  repeatedly  extract  the  cool  urine  with  large 
quantities  of  ether.  Distil  or  evaporate  the  ether.  Para-aminophenol  usually 
appears  as  a  reddish  or  brownish  oil.  An  aqueous  solution  of  this  substance  will 
give  the  indophenol  test. 

PHENACETINE 

Phenacetine,  or  p-aceto-phenetidine,  crystallizes  in  shining  leaflets,  which  are 

without  color,  odor  or  taste,  and  melts  at  134  to  135°.     Phenacetine  is  soluble  in 

■NTXT     r^n,     /-XT    about  1400  parts  of  cold  water,  70  parts  of  boiling  water, 

J-N-tl \^U -1^X13  ,-111  ll-T  111- 

I  16    parts    of   alcohol    and    freely    soluble    in    ether   and 

C  chloroform.     Its  solutions  are  neutral.     Concentrated  sul- 

-/\  phuric   acid   dissolves   it   without   color.     Phenacetine  is 

HC      CH 

I        II  very  closely  related  to  acetaniHde  but  does  not  give  the 

HC      CH  phenylisocyanide  test. 

%/  Preparation. — The    gradual    addition    of    crystallized 

C  phenol  to  cold  dilute  nitric  acid  (sp.  gr.  i.ii  =  17.5  per 

Ip  Tj  cent.)    results  in   the  formation  of  a  mixture  of  o-  and 

p-nitro-phenol.  Since  the  ortho-compound  is  volatile  with 
steam,  complete  separation  of  the  two  products  is  possible  by  steam  distilla- 
tion.    The  residual  p-nitro-phenol  is  converted  into  its  sodium  salt  which  is 


NON-VOLATILE   POISONS 


71 


heated  in  sealed  tube  with  ethyl  bromide  and  thus  changed  to  p-nitro-phenetol. 
The  latter  is  reduced  by  means  of  nascent  hydrogen  from  tin  and  hydrochloric 
acid  to  p-amino-phenetol,  or  p-phenetidine,  which  is  then  boiled  with  glacial 
acetic  acid  and  converted  into  aceto-p-phcnetidine,  or  phenacetine: 


OH 

I 
C 

/\ 
HC      CH 

I       II 

HC      CH 

\/ 

C 


iOH     HiONa 


H 

HOi— NO2 


Phenol 


c 

/\ 

HC      CH 

I       it 
HC      CH 

\/ 
C 

NO2 


p-Nitro- 
phenol 


OC2HB 
I 

C 

/\ 

HC      CH 

HC      CH 

\/ 
C 


HNiH    ; 

IHOI.CO.CHs 

p-Phenetidine 


ONa     Br.CiHj      qCjHj 

'    I 

C 

/\ 
HC      CH 


C 

/\ 
HC      CH 

I       II 
HC      CH 

\/ 
C 

NO2 


HC      CH 

\/ 
C 


NO2 
2H   4H 


Na  salt  of  p- 
nitro-phenol 


p-Nitro- 
phenetol 


OC2H6 
I 

c 

^\ 

HC      CH 

I       II 
HC      CH 

\/ 
G 


NH.CO.CH3 

Phenacetine 


Detection  of  Phenacetine 

The  extraction  of  phenacetine  by  ether  or  chloroform  from 
an  aqueous  tartaric  acid  solution  is  complete. 

1.  Oxidation  Test. — Boil  phenacetine  for  several  minutes 
with  3  cc.  of  concentrated  hydrochloric  acid.  Dilute  with 
10  cc.  of  water  and  filter  when  cold.  A  few  drops  of  chromic 
acid  solution  added  to  the  filtrate  will  gradually  produce  a 
ruby  red  color.  Strong  chlorine  water  may  be  substituted 
for  chromic  acid, 

2.  Indophenol  Test. — Boil  phenacetine  i  or  2  minutes  with 
about  2  cc.  of  concentrated  hydrochloric  acid.  Dilute  with 
water  and  add  a  few  cc.  of  aqueous  carboUc  acid  solution. 
Filter  the  solution  when  cold.  If  a  few  drops  of  freshly  pre- 
pared calcium  hypochlorite  solution  are  added,  the  filtrate  will 


72  DETECTION   OF   POISONS 

have  a  fine  carmine  red  color.  Addition  of  ammonium  hydrox- 
ide solution  in  excess  will  change  this  color  to  violet-blue. 
Freshly  prepared  chlorine  water,  or  3  per  cent,  chromic  acid 
solution,  may  be  substituted  for  hypochlorite  solution  as  an 
oxidizing  agent. 

3.  Autenrieth-HinsbergTest.^ — (a)  WithDilute  Nitric  Acid. 
Heat  phenacetine  to  boiling  with  a  few  cc.  of  dilute  nitric 
acid  (10  to  12  per  cent.).  It  is  soluble  and  gives  an  intense 
yellow  to  orange-red  color.  As  the  solution  cools,  if  sufficiently 
concentrated,  nitro-phenacetine^  will  crystallize  in  long,  yellow 
needles  which  melt  at  103°.  This  test  is  delicate,  and  char- 
acteristic of  phenacetine,  especially  when  nitro-phenacetine  can 
be  obtained  in  crystals  and  its  melting  point  determined.  It 
serves  to  distinguish  phenacetine  from  acetanilide  and  anti- 
pyrine,  both  of  which  give  colorless  solutions  when  warmed  with 
dilute  nitric  acid. 

(&)  With  Concentrated  Nitric  Acid. — A  few  drops  of  con- 
centrated nitric  acid  poured  upon  phenacetine  produce  a  yellow 
to  orange-red  color.  Part  of  the  phenacetine  is  dissolved  with 
the  same  color  and  heat  completes  the  solution.  Nitro-phen- 
acetine crystalHzes  as  the  solution  cools. 

SALICYLIC  ACm 

COOH  Salicylic  acid,  or  ortho-oxy-benzoic  acid,  crystallizes  in  long, 

I  white  needles  soluble  in  about  500  parts  of  cold  and  in  15 

C  parts  of  boiling  water;   and  freely  soluble  in  alcohol,  ether, 

if^\  chloroform  and  carbon  disulphide.     It  has  a  peculiar  taste 

1  II  which  is  sweetish,  acidulous  and  rather  acrid.  It  melts  at  157°. 
HC      CH  Heated  carefully,  salicylic  acid  wiU  sublime  in  fine  needles 

%/  without  decomposition.     A  little  of  the  acid  may  show  this 

^  behavior  even  upon  the  water-bath.    If  heated  quickly,  salicylic 

acid  is  decomposed  in  part  into  phenol  and  carbon  dioxide. 
.COOH 
C6H4  =  CeHs-OH  +  CO2. 

\0H 

^  Archiv  der  Pharmacie  229,  456  (1891). 

2  The  structural  formula  of  mono-nitro-phenacetine  is  as  follows : 

/OC2H5  I 

CeHs— NO2  3. 

^NHCCaHsO)  4 


NON-VOLATILE    POISONS  73 

Concentrated  sulphuric  acid  dissolves  pure  salicylic  acid  without  color  and 
without  decomposition.  The  lead  and  silver  salts  of  this  acid  are  soluble  in  water 
with  difficulty.  Consequently  lead  acetate  will  precipitate  lead  salicylate, 
/COO. 


C0H4  I  Pb,  from  neutral  solutions.     This  salt  is  white,  crystalline  and 

^OH 

soluble  in  hot  water.     It  crystallizes  unchanged  as  the  hot  solution  cools.     Silver 
nitrate  precipitates  white  silver  salicylate. 

R.  Schmitt's  Method  of  Preparation. — Dry  sodium  phenolate  is  kept  cool  and 
saturated  in  an  autoclave  under  pressure  with  carbon  dioxide  (a).  The  sodium 
phenyl-carbonate  undergoes  molecular  rearrangement,  when  heated  at  120-130°, 
and  becomes  isomeric  sodium  salicylate  (/S) : 

^O       ONa  /ONa  ^^^^^^^^^  ^OH  (x) 

(a)  C         +1         =  C  =  0  (/8)  rearrangement    C6H4 

^O        CcHe  ^OCeHs  ^^^^^^  ^COONa     (2) 

Sodium  phenyl-  Sodium  salicyl- 

carbonate  ate 

Detection  of  Salicylic  Acid 

1.  Ferric  Chloride  Test. — Addition  of  ferric  chloride  solution 
to  a  solution  of  salicylic  acid  or  salicylates  produces  a  blue-violet 
color.  If  the  solution  is  very  dilute,  the  color  is  more  of  a  red- 
violet.  Hydrochloric  acid  changes  the  violet  color  to  yellow. 
An  excess  of  the  reagent  affects  the  delicacy  of  the  test. 

This  test  fails  in  presence  of  mineral  acids,  caustic  alkalies  and  alkaline 
carbonates. 

2.  Millon's  Test. — If  an  aqueous  saHcylic  acid  solution  is 
warmed  with  Millon's  reagent,  a  deep  red  color  will  appear. 

3.  Bromine  Water  Test. — This  reagent  in  excess  produces  a 
yellowish  white,  crystalline  precipitate  even  with  very  dilute 
salicyhc  acid  solutions.  The  compound  thus  formed  is  tri- 
bromo-phenyl  hypobromite  (see  page  28). 

OBr 

C 

/OH  ^^ 

/^"-  BrC      CBr 

C6H4  +  4Br2  =  CO2  +4HBr  +  |        || 

^COOH  ^%./^^ 

C 
Br 

4.  Melting-Point  Test. — If  the  quantity  of  saHcylic  acid  is 


74  DETECTION   OF  POISONS 

not  too  small,  dissolve  the  ether  residue  in  very  little  hot  water, 
shake  the  hot  solution  with  a  Uttle  animal  charcoal  and  filter. 
Cool  the  filtrate,  dry  the  crystals  and  determine  the  melting 
point  (iS7°)- 

Separation  of  Salicylic  Acid  from  Simple  Phenols 

When  phenols  like  carboHc  acid  or  the  cresols  are  present, 
the  above  tests  from  i  to  3  prove  nothing  as  far  as  salicylic 
acid  is  concerned.  If  these  compounds  are  present,  add  suffi- 
cient sodium  carbonate  solution  to  render  the  ether  residue 
alkaline  and  extract  the  solution  with  ether.  This  solvent 
will  take  up  the  phenols  and  salicylic  acid  will  remain  in  the 
water  as  the  sodium  salt.  Withdraw  the  aqueous  solution 
from  the  separating  funnel,  acidify  with  dilute  hydrochloric 
or  sulphuric  acid  and  extract  saHcylic  acid  with  ether. 

Directions  are  given  elsewhere  (see  page  243)  for  the  detection 
of  salicylic  acid  in  beer,  milk,  urine,  fruit  juices,  meat  and  meat 
preparations,  as  well  as  in  maltol. 

Quantitative  Estimation  of  Salicylic  Acid  as  Tribromo-phenyl  Hypo- 

bromite 

Place  the  aqueous  solution  of  salicylic  acid  in  a  glass-stop- 
pered flask,  add  an  excess  of  saturated  bromine  water  and  shake. 
The  acid  is  completely  precipitated  as  tribromo-phenyl  hypo- 
bromite.  At  the  end  the  solution  should  be  reddish  brown. 
It  should  stand  12-24  hours  and  be  shaken  frequently.  Collect 
the  precipitate  of  tribromo-phenyl  hypobromite  in  a  weighed 
Gooch  crucible  and  dry  to  constant  weight  in  a  vacuum  desic- 
cator over  sulphuric  acid.  The  quantity  of  salicylic  acid  may 
be  calculated  from  the  weight  of  precipitate  as  follows: 

C6H2Br40:C7H603  =  Wt.  of  precipitate  :  x 
409.86       138.05  obtained 

Detection  of  Salicylic  Acid  in  Urine^ 

Salicylic  acid  forms  a  conjugate  with  glycocoll,  supplied  by  the  organism, 
and  is  changed  in  the  human  body,  in  part  at  least,  into  salicyluric  acid, 

1  To  study  the  behavior  of  saUcyUc  acid  in  this  connection,  take  i  to  1.5  grams 
of  sodium  salicylate  at  night  in  the  course  of  several  hours  and  examine,  as  de- 
Scribed,  the  urine  passed  in  the  next  1 2  hours. 


NON-VOLATILE   POISONS  76 

/OH  (i) 


CoH^C 

V0.NH.CH2.COOH        (2) 

This  is  eliminated  in  urine  with  unaltered  salicylic  acid.  Such  urine  gives  a 
violet  color  with  ferric  chloride  solution.  Both  saUcylic  and  sahcyluric  acids 
give  this  test.  To  decompose  sahcyluric  acid  into  its  constituents,  heat  the  acid 
half  an  hour  with  fuming  hydrochloric  acid  under  a  return-condenser. 

To  isolate  unchanged  salicyHc  acid,  acidify  500  to  1000  cc.  of  urine  with  hydro- 
chloric acid  and  repeatedly  extract  with  ether.  Remove  the  ether  from  the 
aqueous  solution  in  a  separating  funnel  and  shake  vigorously  with  excess  of 
sodium  carbonate  solution.  Salicylic  acid  passes  into  the  aqueous  solution. 
Withdraw  the  aqueous  solution,  which  is  alkahne,  acidify  with  dilute  hydro- 
chloric acid  and  extract  with  ether  which  upon  evaporation  usually  deposits 
the  acid  in  a  crystalUne  condition.  Purify  the  residue  by  recrystallization  from 
water,  using  animal  charcoal  to  remove  color.  Sahcylic  acid  is  rapidly  taken  up 
by  all  mucous  surfaces  and  quickly  absorbed.  Elimination  by  way  of  the  urine 
usually  begins  within  the  first  half  hour  and  is  complete  in  3  days. 

VERONAL 

Veronal,  C-diethyl-barbituric  acid,  C-diethyl-malonyl-urea,  C8H12O3N2, 
crystallizes  from  hot  water  in  large,  colorless,  spear-shaped  crystals  melting  at 
CaHfiv       /CO — NH\  191°  (corrected).     It  is  soluble  in  146-147  parts  of 

y>C<('  ^CO    water  at  20°  and  in  15  parts  at  100°.     Veronal  is  also 

C2H6  CO     NH  freely  soluble  in  hot  alcohol  and  in  acetone.     It  dis- 

solves with  difiiculty  in  cold  ether.  An  aqueous  veronal  solution  has  a  bitter 
taste  and  shows  a  very  faint  acid  reaction  with  sensitive  blue  Utmus  paper. 
Veronal  readily  dissolves  in  caustic  alkalies,  ammonia  and  in  calcium  or  barium 
hydroxide  solution.  From  such  solutions,  provided  they  are  not  too  dilute, 
acids  reprecipitate  veronal  in  a  crystalhne  condition.  Of  the  veronal  salts  the 
sodium  salt,  C8Hii06N2Na,  crystaUizes  best.  It  may  be  prepared  by  dissolving 
veronal  in  the  calculated  quantity  of  caustic  soda  solution  free  from  carbonate, 
and  then  evaporating  this  solution  with  exclusion  of  carbon  dioxide,  or  adding 
alcohol  until  turbidity  appears.  In  both  cases  the  sodium  salt  of  veronal  sepa- 
rates as  splendid  shining  crystals. 

Preparation  by  E.  Fischer  and  A.  Dilthey^ 

(a)  From  diethyl-ethylmalonate  by  condensation  with  urea  in  presence  of 
sodium  ethylate: 

NaiOCzHs 
CjHs  CO— lOCaSis  HInIh  C2H6  CO— N— Na 

)>C(  ^^   ^  \     /  \^^   _^  3C2H5.OH. 


C2H5  CO— OCzHb  HNH  C2H5  CO— NH 

Diethyl-ethylmaionate       Urea  Na  salt  of  veronal 

^  Annalen  der  Chemie  und  Pharmazie,  335,  334  (1904). 


76  DETECTION    OF   POISONS 

Dissolve  metallic  sodium  (3  2  parts)  in  absolute  alcohol  (600  parts)  and  when 
cold  add  diethyl-ethylmalonate  (100  parts).  Dissolve  in  this  mixture  with  heat 
finely  powdered  urea  (40  parts).  Heat  4-5  hours  in  an  autoclave  at  105-108°. 
The  sodium  salt  of  veronal  is  precipitated  even  from  the  hot  solution  as  a  color- 
less, crystalline  mass.  Cool,  filter  with  suction  and  wash  with  alcohol.  Dis- 
solve the  crystals  in  water  and  acidify  with  concentrated  hydrochloric  acid. 
Veronal  thus  precipitated  is  pure  when  recrystallized  from  water. 

(b)  From  diethyl-malonyl  chloride  by  condensation  with  urea: 


CaHs  CO— :C1     H:NH  C2H5  CO— NH 

^c/        ^co = 2HC1  +   y^^     /CO 

C2H5  CO— iCl  "  HINH  C2H6  CO— NH 


Heat  diethyl-malonyl  chloride  (3  parts)  on  the  water-bath  for  20  hours  with 
finely  powdered,  dry  urea  (2  parts).  Considerable  hydrochloric  acid  is  given 
off  toward  the  end  and  a  solid  mass  finally  remains  which  yields  pure  veronal 
upon  crystallization  from  hot  water.  The  yield  is  70  per  cent,  of  the  theoretical 
amount. 

Physiological  Action. — Veronal  does  not  cause  decomposition  of  the  blood  and 
in  the  usual  medicinal  doses  (0.5-1  gram)  does  not  appear  to  act  strongly  upon 
the  heart.  Cumulative  action  has  been  noted  only  in  rare  instances.  In  large 
doses,  however,  veronal  may  cause  serious  intoxication  with  fatal  termination. 
Death  resulted  in  the  case  of  a  man  in  Holzminden  who  had  taken  10  grams  of 
veronal  by  accident.  There  are  also  on  record  two  other  suicidal  cases,  one  from 
II  grams  and  the  other  from  15  grams  of  this  hypnotic.  There  was  loss  of  con- 
sciousness and  contraction  of  the  pupils  in  the  second  case.  Atropine  caused 
dilatation  of  the  pupils  but  otherwise  was  of  no  avail.     Death  ensued  in  20  hours. 


Detection  of  Veronal 

In  examining  cadaveric  material  (liver,  spleen  and  kidneys) 
from  a  man,  who  had  taken  veronal,  thinking  it  was  a  remedy 
for  tape-worm  (kamala),  G.  and  H.  Frerichs^  isolated  small 
quantities  of  this  drug.  Following  the  Stas-Otto  process,  they 
extracted  the  aqueous  tartaric  acid  solution  with  ether  and 
evaporated  the  ether  extract.  They  recrystallized  the  residue 
from  a  little  hot  water,  using  animal  charcoal  to  remove  color, 
and  identified  the  crystals  obtained  as  veronal  by  the  following 
tests : 

1.  The  aqueous  solution  of  the  crystals  had  a  faintly  acid 
reaction. 

2.  The  crystals  were  soluble  in  sodium  hydroxide  solution 

^  Archiv  der  Pharmazie  244,  86-90  (1906). 


NON-VOLATILE   POISONS  77 

and  ammonia  and  were  reprecipitated  when  the  solutions  were 
acidified  with  dilute  hydrochloric  acid. 

3.  The  melting-point  of  the  crystals  was  187-188'^.  On 
mixing  the  substance  with  pure  veronal,  they  obtained  the  same 
melting-point.  ^ 

4.  Presence  of  nitrogen  was  shown  by  fusing  the  crystals  in  a 
dry  test-tube  with  metallic  sodium,  coohng  the  melt,  dissolving 
in  water  and  testing  for  sodium  cyanide  by  the  Prussian  blue 
reaction  (see  page  22). 

5.  The  crystals  were  heated  in  a  dry  test-tube  and  sublimed. 
They  were  then  compared  with  crystals  known  to  be  pure 
veronal  and  found  identical. 

Detection  of  Veronal  in  Urine 

E.  Fischer  and  J.  v.  Mering,^  and  also  B.  Molle  and  H.  Kleist,^ 
have  found  that  most  of  the  veronal  leaves  the  human  body 
unchanged  and  is  present  in  the  urine  to  the  extent  of  70-90 
per  cent.  Consequently  in  veronal  poisoning  the  urine  should 
be  examined  first.  Concentrate  a  considerable  quantity  on 
the  water-bath^  to  one-fifth  its  volume  and  extract  several 
times  with  ether,  using  a  large  volume  at  each  extraction  because 
veronal  is  not  very  soluble  in  this  solvent.  The  residue  left 
after  distilling  the  ether  is  usually  quite  dark  in  color.  Dissolve 
in  as  little  hot  water  as  possible,  boil  the  solution  15  minutes 
with  animal  charcoal  and  filter.  Cool  the  nearly  colorless 
filtrate  with  ice  and  veronal  will  crystalHze  in  colorless  needles 
melting  at  191°  (corrected). 

E.  Fischer  and  v.  Mering  recovered  from  5  daj's  urine,  after  administration 
of  4  grams  of  veronal  during  2  days,  2.49  grams  =  62  per  cent,  of  the  quantity 
used.     The  method  therefore  is  not  absolutely  quantitative  nor  is  the  elimination 

^  A  mixture  of  two  organic  substances,  having  the  same  melting-point  but  not 
being  identical,  will  show  a  melting-point  lower  than  that  of  either  substance 
taken  by  itself.  But  obviously  a  mixture  of  identical  substances  will  show  no 
depression  of  melting-point  whatever. 

^  Die  Therapie  der  Gegenwart  45,  1904. 

'  Arcliiv  der  Pharmazie  242,  401  (1904). 

*  Fischer  and  v.  Mering  evaporated  the  urine  under  diminished  pressure. 


78  DETECTION   OF   POISONS 

of  veronal   complete   after   5  days.     The  crystals  obtained   should  be  proved 
conclusively  to  be  veronal  by  the  tests  described. 

Molle  and  Kleist  first  add  lead  acetate  solution  to  the  urine  as  long  as  it  causes 
a  precipitate,  filter,  remove  lead  from  the  filtrate  by  hydrogen  sulphide  and  filter 
from  lead  sulphide.  Hydrogen  sulphide  is  expelled  with  heat  and  the  urine, 
after  dilution  with  twice  its  volume  of  water,  is  boiled  with  animal  charcoal. 
The  filtrate,  after  concentration  on  the  water-bath  to  a  small  volume,  is  cooled, 
saturated  with  sodium  chloride  and  extracted  3  times  with  ether.  The 
filtered  ether  solution  on  distillation  leaves  nearly  pure  veronal. 

ANTIPYRINE 

Antipyrine,  or  i-phenyl-2, 3-dimethyl-isopyrazolone,  C11H12ON2,  forms 
monoclinic,  tabular  crystals  having  a  faintly  bitter  taste  and  melting  at  113°. 

One  part  of  antipyrine  is  soluble  in  less  than  i 
part  of  cold  water,  in  about  i  part  of  alcohol,  i 
part  of  chloroform  and  in  about  50  parts  of 
ether.  An  aqueous  antipyrine  solution  has  a 
neutral  reaction,  although  this  compound  is  a 
base  and  forms  crystaUizable  salts  with  acids. 
Preparation.  Antipyrine  is  formed  directly  by  heating  ;8-phenyl-methyl- 
hydrazine  and  aceto-acetic  ester: 

CH3— CiO     Hi  N— CH3  (3)  CH3— C  —  N— CH3   (2) 


(3) 

CH3 

— C  —  N- 

-CH3 

(2) 

HC        N- 

\/ 
C 
0 

-CeHs 

(i) 

HCiH:     ;HiN— CeHs    =  HC        N— CeHsCi) +H2O  +  C2H5.OH. 

\'      i      ': ;  \/ 

C—0  C2H6:  c 

o  o 

Detection  of  Antipyrine 

Ether  extracts  only  small  quantities  of  antipyrine  from  a 
solution  containing  much  tartaric  acid.  Ether,  or  better 
chloroform,  extracts  by  far  the  greater  part  of  the  antipyrine 
when  the  solution  has  been  made  alkaline.  Antipyrine  differs 
from  most  alkaloids  in  being  more  soluble  in  water.  To  detect 
antipyrine,  dissolve  the  residue  left  on  evaporating  the  ether 
solution  in  a  Httle  water,  filter  and  apply  the  following  tests: 

1.  Ferric  Chloride  Test. — Add  i  or  2  drops  of  ferric  chloride 
solution  to  an  aqueous  antipyrine  solution.  It  will  produce 
a  deep  red  color  which  can  be  seen  even  in  a  dilution  of  i  : 
100,000. 

2.  Tannic  Acid  Test. — Tannic  acid  solution  produces  an 
abundant,  white  precipitate,  when  added  to  an  aqueous  anti- 
pyrine solution. 


NON-VOLATILE    POISONS  79 

3.  Fuming  Nitric  Acid  Test. — Dissolve  antipyrinc  in  a  few 
drops  of  water  and  add  i  or  2  drops  of  fuming  nitric  acid.  The 
solution  will  be  green.  If  this  solution  is  heated  to  boiling, 
another  drop  of  nitirc  acid  will  produce  a  red  color.  Two  cc. 
of  antipyrine  solution  (i  :  200)  will  give  this  test  distinctly. 

4.  Nitroso-antipyrine  Test. — Add  a  few  drops  of  potassium 
or  sodium  nitrite  solution  to  an  aqueous  antipyrine  solution 
and  then  dilute  sulphuric  acid.  A  green  or  blue  color  will 
appear.  A  few  drops  of  acetic  acid  may  be  substituted  for 
sulphuric  acid  but  the  solution  must  be  heated.  If  the  anti- 
pyrine solution  has  been  concentrated,  green  crystals  of  nitroso- 
antipyrine  (CiiHii(N0)0N2)  will  separate  after  some  time. 

Detection  of  Antipyrine  in  Urine 

Part  of  the  antipyrine  passes  unchanged  into  the  urine  but  some  is  also  present 
as  oxy-antipyrine-glycuronic  acid,  a  direct  test  for  which  may  be  made  by  means 
of  ferric  chloride  solution.  When  the  urine  is  highly  colored,  render  a  consider- 
able quantity  alkaline  with  sodium  hydroxide  solution  or  ammonia  and  extract 
with  chloroform.  Dissolve  the  residue  left  by  chloroform  in  a  little  water  and 
test  for  antipyrine  with  ferric  chloride  solution  and  with  fuming  nitric  acid. 

CAFFEINE 

Caffeine  (theine)  or  i,3,7-trimethyl-2,6-dioxy-purine  (C8H10O2X4.H2O), 
crystallizes  in  white,  shining  needles.     It  is  soluble  in  80  parts  of  water,  giving 

(1)  CH N 00(6')  ^  colorless  solution  with  a  neutral  reaction  and 

I       I  /CH3(7)       3,  faint,  bitter  taste.     Caffeine  is  quite  easily 

(2)  OC     C — N<('  soluble  in  hot  water  (1:2).     Jt  requires  for 

I       II        ^^-^  solution   nearly   50   parts   of   alcohol,  only  9 

^^'         *  parts  of  chloroform  and  is  only  slightly  soluble 

in  ether.  In  crystallizing  from  hot  water  caffeine  combines  with  i  molecule  of 
water,  a  part  of  which  it  loses  upon  exposure  to  air  and  all  when  dried  at  100°. 
Caffeine  is  only  very  sHghtly  soluble  in  absolute  alcohol,  benzene  and  petroleum 
ether.  It  melts  at  230°,  but  somewhat  above  100°  begins  to  volatilize  in  small 
quantity  and  at  180°  to  sublime  without  leaving  a  residue.  Concentrated  sul- 
phuric and  nitric  acids  dissolve  it  without  color.  Caffeine  is  a  ven,"  weak  base 
and  its  salts  are  decomposed  by  water.  Therefore,  caffeine  can  be  extracted  at 
least  partially  by  ether,  or  better  by  chloroform,  from  an  aqueous  tartaric  acid 
solution.  The  relation  existing  between  caffeine  and  uric  acid  is  quite  apparent 
when  the  products,  formed  by  oxidizing  these  two  substances  with  potassium 
chlorate  and  hydrochloric  acid,  are  compared.  Oxidation  of  uric  acid  \-ields 
alloxan  and  urea;  cafl'eine  gives  dimethji-alloxan  and  monomethyl-urea. 


80 


CHs— N— CO 

I       I 

oc   c 

I    II 

CH3— N— C 

Caffeine 


DETECTION   OF  POISONS 

CH3— N— CO       NH— CH3 

I       I  I 

OC     CO  +  CO 


— N^CH3  +  (H2O)  +  02  = 


CH3— N— CO       NH2 

Dimethyl-        Monomethyl- 
alloxan  urea 


Fate  of  Caffeine  in  Hiiman  Metabolism. — Only  a  very  small  part  of  the  cafEeine 
taken  into  the  body  passes  through  unchanged  and  appears  in  the  urine.  About 
10  per  cent,  appears  in  the  urine  as  decomposition  products.  The  remainder 
may  be  changed  into  normal  end-products  of  human  metabolism.  Most  of  the 
nitrogen  of  caffeine  is  eliminated  as  urea.  A  very  important  fact  is  the  cleavage 
of  methyl  groups  with  formation  of  the  first  decomposition  products  of  caffeine, 
namely,  dimethyl-  and  monomethyl-xanthines.  Of  the  monomethyl-xanthines, 
7-monomethyl-xanthine  is  formed  especially.  Of  the  dimethyl-xanthines, 
paraxanthine  =  1,7-dimethyl-xanthine  is  found.  Both  of  these  compounds 
appear  in  urine  after  administration  of  caffeine.  Paraxanthine  is  isomeric 
with  theophylline,  or  1,3-dimethyl-xan thine,  and  with  theobromine,  or 
3 , 7-dimethyl-xanthine . 

The  structural  formulae  of  these  cleavage-products  of  caffeine  in  animal  meta^ 
holism  are  as  follows: 


HN— CO 

I       I 
OC     C— N 


I        II  /A 

HN— C— N^ 

7-M  ethyl  -xanthine 


CHa  (7) 
CH 


(i)  CH3.N— CO 

OC     C— NH 1 

I    II    > 

(3)  CH3.N— C— N^ 
Theophylline 


CH 


(i)  CH3.N— CO 


OC     C— N.CH3  (7) 

HN— C— N^ 
Paraxanthine 


HN— CO 

OC     C— N.CH3  (7) 

^JCH 
(3)  CH3.N— C— N^ 

Theobromine 


r/^ 


Detection  of  Caffeine 

Ether  will  extract  more  caffeine  from  an  aqueous  alkaline 
solution  than  from  an  aqueous  tartaric  acid  solution.  Since 
caffeine  dissolves  with  some  difficulty  in  ether,  but  more  easily 
in  chloroform,  the  latter  solvent  is  usually  employed  after  the 
solution  has  been  made  alkaline  with  ammonia.  After  dis- 
tillation of  solvent,  caffeine  appears  in  concentric  clusters  of 
long,  shining  needles.  In  an  analysis  by  the  Stas-Otto  method 
caffeine  will  appear  in  all  three  extracts. 


NON-VOLATILE    POISONS  81 

1.  Oxidation  Test. — Pour  a  few  cc.  of  saturated  chlorine 
water ^  over  caffeine  and  evaporate  the  solution  to  dryness  upon 
the  water-bath.  A  reddish  brown  residue  will  remain.  If  a 
few  drops  of  ammonium  hydroxide  solution  are  added,  a  fine 
purple  red  color  will  immediately  appear.  This  test  may  be 
made  by  covering  the  dish  containing  the  residue  with  a  glass 
plate  moistened  with  a  drop  of  strong  ammonia.  Or  two 
matched  watch  glasses  may  be  used,  the  material  containing 
caffeine  being  evaporated  to  dryness  with  chlorine  water  upon 
one  glass  which  is  then  placed  for  a  short  time  upon  the  other 
glass  containing  a  drop  of  strong  ammonia. 

This  test,  known  as  the  murexide  reaction,  is  also  given  by 
xanthine,  theobromine,  i-  and  y-monomethyl-xanthine  and 
paraxanthine,  especially  jf  made  as  described  by  E.  Fischer.  ^ 
Heat  the  material  to  boiling  in  a  test-tube  with  strong  chlorine 
water,  or  with  hydrochloric  acid  and  a  little  potassium  chlorate, 
evaporate  the  liquid  to  dryness  in  a  dish  and  moisten  the  residue 
with  ammonia. 

2.  Tannic  Acid  Test. — This  reagent,  added  to  an  aqueous 
caffeine  solution,  causes  a  heavy  white  precipitate  which  is 
soluble  in  an  excess  of  the  acid.  This  test  is  not  characteristic 
of  caffeine. 

B.  Examination  of  Ether  Extract  of  Alkaline  Solution 
(Most  of  the  alkaloids  appear  here) 
Add  enough  sodium  hydroxide  solution  to  the  acid  solution 
separated  from  ether  to  make  it  strongly  alkaUne.  The  alkali 
will  liberate  alkaloids  from  their  salts  and  combine  with  mor- 
phine and  apomorphine,  if  present.  Thoroughly  extract  this 
alkaline  solution  with  about  the  same  quantity  of  ether. 
This  solvent  will  dissolve  all  alkaloids  except  morphine, 
apomorphine  and  narceine.  Separate  the  ether  from  the 
aqueous  solution  and  again  extract  with  a  fresh  quantity 
of   ether.     In    certain    cases   3    or   4    such    extractions   may 

^  A  convenient  method  of  preparing  a  saturated,  aqueous  chlorine  solution  is  to 
heat  potassium  chlorate  with  hydrochloric  acid  and  pass  the  chlorine  into  a  small 
quantity  of  water. 

^Berichte  der  Deutschen  chemischen  Gesellschaft  30,  2236  (1S97). 


82 


DETECTION   OF   POISONS 


be  required.  Pour  the  ether  extracts  into  a  dry  flask,  stop- 
per loosely  and  set  aside  for  i  or  2  hours.  A  few  drops 
of  water  always  settle  to  the  bottom  of  the  flask.  Carefully 
decant  the  ether  and  pour  through  a  dry  filter.  Evaporate  the 
filtrate  with  gentle  heat  in  a  glass  dish  (8  to  10  cm.  in  diame- 
ter). Let  the  last  part  of  the  ether  solution  evaporate  spon- 
taneously. If  small  globules  having  a  strong  odor  appear,  the 
residue  must  be  examined  for  coniine  and  nicotine.  If  there  is 
no  trace  of  these  volatile  alkaloids,  gently  heat  the  residue 
upon  the  water-bath  to  expel  water  left  by  evaporation  of  the 
ether.  Remove  the  dish  from  the  water-bath  as  soon  as  this 
has  been  accomplished.  It  is  not  advisable  to  heat  the  residue 
too  long,  as  it  tends  to  become  viscous.  This  residue,  obtained 
by  extracting  the  alkaline  solution  with  ether,  may  contain 
any  alkaloid  except  morphine,  apomorphine  and  narceine.  It 
should  be  examined  for 


Coniine 

Brucine 

Codeine 

Qiunine 

Nicotine 

Atropine 

Narcotine 

Caffeine 

Anilinfi 

Scopolamine 

Hydrastine 

Antipyrine 

Veratrine 

Cocaine 

Pilocarpine 

Pyramidone. 

Strychnine 

Physostigmine 

First,  note  the  general  appearance  of  the  residue  and  then 
examine  with  the  microscope.  Taste  it  cautiously.  Certain 
alkaloids  may  be  recognized  beforehand  by  this  test.  Special 
tests  should  then  be  made  at  once.  The  various  alkaloids 
appear  in  the  residue  as  follows : 

Strychnine. — Fine  needles  having  an  exceedingly  bitter  taste. 

Brucine. — Usually  a  white,  amorphous  powder  having  a 
very  bitter  taste. 

Veratrine. — Usually  an  amorphous  powder  having  a  sharp, 
burning  taste. 

Atropine  and  Quinine. — A  varnish  which  is  resinous  and 
sticky.     Rarely  crystalline  or  in  the  form  of  a  powder. 

Codeine. — A  thick,  viscous  syrup  which  after  a  time  becomes 
solid,  especially  if  stirred  with  a  glass  rod,  and  frequently 
crystalline. 


NON-VOLATILE   POISONS  83 

Caffeine. — ^Long,  silky  needles  having  a  faintly  bitter  taste. 
These  are  frequently  concentrically  arranged. 

Antipyrine. — A  syrup  which  gradually  becomes  crystalline, 
especially  if  stirred.  It  has  a  mild,  bitter  taste  and  dissolves 
very  easily  in  water.  ^ 

Pyramidone. — Usually  as  fine  needles  which  have  a  faintly 
bitter  taste.     It  is  easily  soluble  in  water. 

Frequently  ether  leaves  only  a  slight,  tasteless  residue.  In 
that  case  alkaloids  are  absent.  Such  residues  often  consist 
of  fat,  resinous  matter,  or  traces  of  nitrogenous  substances 
(Peptones  and  their  cleavage  products?  Creatinine?).  Parts 
of  a  cadaver,  even  when  quite  fresh,  usually  give  small  residues 
at  this  point.  Alkaloids  may  be  absent  and  every  step  in  the 
process  may  have  been  performed  with  the  greatest  care.  To 
be  quite  sure  that  alkaloids  are  absent,  dissolve  a  portion  of  the 
residue  in  water  containing  a  drop  of  dilute  hydrochloric  acid. 
Filter,  if  necessary,  and  distribute  this  solution  upon  several 
watch-glasses.     Test  with  the  following  alkaloidal  reagents: 

Mercuric  Chloride  Picric  Acid 

lodo-Potassium  Iodide  Tannic  Acid 

Potassium  Mercuric  Iodide  Phospho-Molybdic  Acid 

Potassium  Bismuthous  Iodide  Phospho-Tungstic  Acid. 

Unless  these  reagents  give  distinct  and  characteristic  pre- 
cipitates, alkaloids  are  absent.  It  is  advisable  in  every  instance 
to  make  this  preliminary  test  for  alkaloids.  Only  a  small  por- 
tion of  material  is  required  and  these  general  reagents  show  even 
traces  of  alkaloids. 

To  exclude  mistakes  and  oversights  in  toxicological  analysis,  dissolve  the  ether 
residue,  should  it  be  very  small,  in  a  few  cc.  of  very  dilute  hydrochloric  acid  (about 
I  per  cent,  of  HCl).  Evaporate  this  solution  upon  the  water-bath  and  dissolve 
the  residue  in  a  little  water.  Inject  this  solution  from  a  h}-podermic  sj-ringe  into 
the  lymph-sac  on  the  back  of  a  small  but  lively  frog.     If  the  frog  shows  no  sign  of 

^  Most  of  the  alkaloids  are  onh'  slightlj-  soluble  in  cold  water.  Some  cannot  be 
detected  satisfactorily  by  purely  chemical  means.  Others  have  no  characteristic 
tests.  Such  substances  should  not  be  selected  for  laboratorj-  practice.  They 
may  cause  beginners  to  think  that  the  experienced  toxicologist  relies  upon  similar 
uncertain  methods  when  he  seeks  to  identifv  an  alkaloid  in  an  actual  analvsis. 


84  DETECTION   OF   POISONS 

poisoning  in  the  course  of  several  hours,  it  is  quite  likely  that  the  residue  does  not 
contain  any  very  poisonous  alkaloid. 

In  making  special  tests  for  alkaloids,  distribute  the  residue 
upon  several  watch  glasses,  using  a  platinum  or  nickel  spatula 
or  a  small  penknife.  Or  dissolve  the  residue  in  a  little  hot 
alcohol,  filter  the  solution,  distribute  upon  watch  glasses  and 
evaporate  at  a  gentle  heat.  R.  Mauch^  dissolves  the  residue 
in  75  per  cent,  aqueous  chloral  hydrate  solution  and  uses  this 
solution  in  testing  for  alkaloids.  (The  details  of  this  method 
will  be  found  on  page  244.) 

Purification  of  the  Alkaloidal  Residue 

If  alkaloids  are  contaminated  with  greasy,  resinous  or  fatty 
substances,  many  of  the  tests  will  either  fail  entirely  or  give 
uncertain  results.  In  this  case  the  residue  must  be  purified 
in  one  of  two  ways. 

1.  Thoroughly  mix  the  residue  with  cold  water  containing 
hydrochloric  acid.  Filter  to  remove  insoluble  matter  (fatty 
or  resinous  substances),  add  sodium  hydroxide  solution  to  the 
filtrate  until  alkahne  and  extract  with  ether.  The  alkaloids 
obtained  by  evaporating  the  solvent  are  usually  quite  pure. 

2.  Or  dissolve  the  residue  in  hot  amyl  alcohol,  extract  this 
solution  with  a  few  cc.  of  very  dilute  sulphuric  acid  and  with- 
draw the  acid  solution  from  the  separating  funnel.  Amyl 
alcohol  will  retain  greasy  and  colored  impurities,  and  the  alka- 
loids will  be  in  the  aqueous  solution  as  sulphates.  Add  sodium 
hydroxide  solution  in  excess  and  extract  with  ether.  This 
method  of  purifying  the  alkaloidal  residue  is  especially  recom- 
mended, when  there  is  considerable  coloring  matter. 

W.  H.  Warren  and  R.  S.  Weiss ^  have  suggested  picrolonic  acid^  as  a  means  of 
purifying  alkaloids.  An  alkaloid  like  strychnine,  whose  picrolonate  is  very  insolu- 
ble, may  be  precipitated  from  aqueous  solution  and  thus  separated  from  other 
substances  which  prevent  purification.     The  precipitated  picrolonate  may  be 

^  Richard  Mauch  (Mittheilungen  aus  dem  Institut  des  Herrn  Prof.  Dr.  E. 
Schaer  in  Strassburg),  "Festgabe  des  Deutschen  Apotheker-Vereins,"  Strassburg, 
1897. 

^  The  Journal  of  Biological  Chemistry,  3,  330  (1907). 

^  For  the  preparation  of  this  reagent,  see  page  313. 


NON-VOLATILE   I'OLSONS  85 

collected  on  a  filter,  washed  with  water  and  then  warmed  with  dilute  sulphuric 
acid  which  discharges  the  bright  yellow  color  of  the  picrolonate  causing  the  alka- 
loid to  pass  into  solution  and  precipitating  pale  yellow  picrolonic  acid.  By 
extracting  with  acetic  ether,  in  which  picrolonic  acid  is  especially  soluble,  the 
aqueous  solution  of  the  alkaloid  is  left  colorless.  Neutralization  with  sodium 
hydroxide  solution  and  extraction  with  ether  will  give  a  very  pure  alkaloid. 

CONIINE 

Coniine,  a-normal-propyl-piperidine,  CsHwN,  occurs  in  all  parts  of  spotted 

hemlock   (Conium   maculatum)   together  with    n-methyl-coniine,  conhydrine, 

Ho  7-coniceine  and  pseudo-conhydrine.     It  is  a  color- 

C  less,    oily,    very    poisonous    liquid    which    becomes 

„  p   \-TT  yellowish  or  brown  in  contact  with  air  and  is  par- 

^i  ^        I     '  tially  resinified.      It  is  slightly  soluble  in  cold  but 

H2C    *CH.CIi2.CH2.CH3  fi'^en  less  soluble  in  hot  water.     Coniine  is  miscible 

\/  with   alcohol,   ether,   chloroform  and  benzene  in  all 

^  proportions.      The  unpleasant,  narcotic  odor  of  this 

alkaloid,  sometimes  said  to  resemble  that  of  mouse 

urine,    is   more  intense   than   the  odor  of  nicotine.     Coniine  as  it  occurs  in 

nature  is  dextro-rotatory,i  [q;]d  =  -[-18.3°,  and  rather  a  strong  base.     Heated 

with  acetic  anhydride,  it  forms  acetyl-coniine : 


CsHieN 
CH3.CO. 


O.CO.CH3    =  CsHieN.CO.CHs  +  CH3.COOH; 


Shaken  with  benzoyl  chloride  and  sodium  hydroxide  solution,  it  forms  benzoyl- 
coniine: 

C^H^.CO  CI   +  -^^^^  =  CsHieN.CO.CeHs  +  NaCl  +  H2O; 
and  with  nitrous  acid  nitroso-coniine: 

^'■^  OH  =  CsHieN.NO  +  U,0. 
All  these  reactions  show  that  coniine  is  a  secondary  base. 

Detection  of  Coniine 

The  alkaloidal  reagents  especially  delicate  with  coniine  are: 
iodo-potassium  iodide  (i:8ooo),  phospho-molj^bdic  acid 
(1:5000),  potassium  mercuric  iodide  (i:8ooo)  and  potassium 
bismuthous  iodide  (1:5000).  Gold  and  platinum  chlorides 
fail  to  precipitate  coniine  when  the  concentration  is  less  than 

1  The  optical  activity  of  this  alkaloid  is  occasioned  by  the  presence  of  the  asym- 
metric carbon  atom  marked  mth  an  asterisk  in  the  structural  formula. 


86  DETECTION   OE   POISONS 

I :  loo;  whereas  they  will  precipitate  nicotine  when  the  concen- 
tration of  the  solution  is  as  low  as  i :  10,000  and  i :  5000. 
When  coniine  is  present,  the  residue  left  by  the  ether  solution 
has'the  characteristic  odor  of  this  alkaloid.  The  two  following 
tests  should  then  be  made: 

1.  Solubility  Test. — Dissolve  a  drop  of  coniine  in  just  enough 
cold  water  to  give  a  clear  solution.  Gently  heat  the  solution 
and  it  will  become  milky,  because  coniine  is  more  easily  soluble 
in  cold  than  in  hot  water.  A  coniine  solution  which  is  milky 
when  hot  becomes  clear  on  cooling.  Aqueous  coniine  solutions 
have  an  alkaline  reaction.  Test  the  solution  with  red  litmus 
paper. 

2.  Crystallization  Tpst. — Put  a  Httle  coniine  upon  a  watch 
glass,  or  glass  slide,  and  add  i  or  2  drops  of  hydrochloric  acid. 
Evaporate  to  dryness  and  coniine  hydrochloride  (CsHnN.HCl) 
will  remain.  Immediately  after  evaporation  examine  this  resi- 
due with  a  microscope  magnifying  about  200  times.  The  color- 
less or  faintly  yellow  crystals  are  needle-Kke,  or  columnar  and 
frequently  grouped  in  star-shaped  clusters.  They  show  the 
play  of  color  characteristic  of  doubly  refractive  substances. 

NICOTINE 

Nicotine,  C10H14N2,  is  a  colorless  hygroscopic  liquid  which  soon  turns  yellow 

and  then  brown  upon  exposure  to  air  and  in  time  becomes  resinous.     It  is  miscible 

with  water  in  all  proportions  (distinction  from  coniine) 

■^  ^         and  freely,  soluble  in  alcohol,  ether,  amyl  a,lcohol,  ben- 

C  L-XI2 L,±l2  ,  ,  ,  ^  ,  .        . 

/<\        I  I  zene  and  petroleum    ether.      Jbther    extracts    nicotine 

HC    /3C — CHa    CH2    from  aqueous  solution.      It  has  a  sharp,  burning  taste 
II        I        \     /  and    strong    odor   of   tobacco    especially   when   warm. 

V     yT  I  Chemically  pure  nicotine  is  said  to  be  almost  inodorous. 

'  N  CHa         ^^^  so-called  tobacco  odor  is  developed  after  the  alka- 

loid has  been  for  some  time  in  contact  with  air.     The 

free  alkaloid  is   strongly  laevo-rotatory,    [ajo   =    —161.55°,  but  its  salts  are 

dextro-rotatory. 
Constitution. — Nicotine  is  a  rather  strong  di-acid,  ditertiary  base  and  forms 

well-crystallized  salts  with  one  or  two  equivalents  of  acid.     Like  ditertiary  bases 

it  combines  with  two  molecules  of  methyl  iodide^  forming  a  di-iodo-methylate, 

'  ^  In  methyl  iodide — as  well  as  in  other  alkyl  haloids — we  have  an  excellent 
means  of  recognizing  the  tertiary  nature  of  a  nitrogen  base.    Like  trimethylam- 


NON-VOLATILE    POISONS  87 

C10HHN2.2CH3I.  Oxidized  wilh  chromic  acid,  nitric  acid  or  potassium  per- 
manganate, nicotine  is  converted  into  nicotinic  acid,  or  /J-carboxy-pyridine. 
Tliis  shows  that  nicotine  is  a  pyridine  derivative  having  a  side-chain  in  the  /3- 
position  with  respect  to  the  pyridine  nitrogen. 

H  H 

C         CH2 — CH2  ^ 

/Nil  /N 

HC   ^C— CH«    CH2  HC      C— COOH 

I        II         \/  I       li 

HC      CH        N  gives  on  oxidation  HC      CH 

\/  I  \/ 

N  CH3  N 

Nicotine  Nicotinic  acid 


This  formula  for  nicotine  proposed  by  Pinner  was  confirmed  several  years  later 
by  Ame  Pictet's  synthesis  of  this  alkaloid. 

Physiological  Action. — Nicotine  is  one  of  the  most  powerful  poisons  and  scarcely 
inferior  to  hydrocyanic  acid  in  toxicity  and  rapidity  of  action.  It  appears  to  be 
toxic  to  all  classes  of  animals.  It  is  absorbed  from  the  tongue,  the  eye  and  the 
rectum  even  in  a  few  seconds  and  from  the  stomach  somewhat  more  slowly. 
Absorption  of  nicotine  is  also  possible  from  the  outer  skin.  Elimination  takes 
place  through  the  lungs  and  kidneys.  In  concentrated  form  nicotine  is  a  local 
irritant,  though,  owing  to  the  rapidity  of  its  toxic  action,  it  does  not  behave  like  a 
true  corrosive  nor  does  it  cause  inflammation  of  the  mucous  lining  of  the  stomach 
even  after  a  lethal  dose.  Nicotine,  after  causing  stimulation  for  a  brief  period, 
then  paralyzes  the  central  nervous  system  and  spinal  cord,  finally  affecting  vari- 
ous organs  such  as  the  heart,  eyes  and  intestinal  tract.  Its  poisonous  influence 
probably  extends  to  all  parts  of  the  brain,  medulla  oblongata  and  spinal  cord. 
Huchard  states  that  nicotine  causes  a  general  convulsion  of  the  circulatory  sys- 
tem which  is  apparent  in  chronic  nicotine  poisoning.  In  chronic  tobacco  poison- 
ing the  general  condition  of  health  is  disturbed  and  quite  frequently  the  eyes  are 
affected.  In  acute  nicotine  poisoning  death  ensues  from  paralysis  of  the  respira- 
tory center.  An  action  upon  the  heart  is  also  always  in  evidence  even  in  non- 
fatal cases. 

ine,  tertiary  cyclic  amines,  as  pyridine  and  quinoline,  also  give  similar  iodo- 
methylates  which  are  ammonium  iodides  with  quinquivalent  nitrogen : 

H  H 

C  C 

CH3\  III  rS' 

CHa^N        +CH3.I  =    pS'llN.I;  HC  CH                      HC  CH 

CH3/                                    ^gV                I  II     +CH3.I=       I  II 

Trimethylamine                                     ^  HC  CH  HC  CH 


C  CH  HC         ( 

I  II     +CH3.I=       I 

C         CH  HC  ( 


N 

PjTidine  /      \ 

H3C        I 


88  DETECTION   OF   POISONS 

Detection  of  Nicotine 

Ether  or  low-boiling  petroleum  ether  will  extract  nicotine 
from  an  aqueous  alkaline  solution.  Spontaneous  evaporation 
of  the  solvent  leaves  the  alkaloid  as  an  oily  liquid  having  the 
odor  of  tobacco  and  a  strong  alkaline  reaction.  General  alka- 
loidal  reagents  will  precipitate  nicotine  from  quite  dilute  solu- 
tions, in  which  respect  this  alkaloid  is  very  different  from  coni- 
ine.  Phospho-molybdic  acid  and  potassium  bismuthous  iodide 
precipitate  nicotine  even  in  a  dilution  of  i  :  40,000;  potas- 
sium mercuric  iodide  in  1:15,000;  gold  chloride  in  1:10,000; 
and  platinum  chloride  in  i :  5000. 

1.  Crystallization  Test. — Evaporate  nicotine  on  a  watch 
glass  with  a  few  drops  of  concentrated  hydrochloric  acid.  This 
will  yield  a  yellow,  varnish-like  residue  which  microscopic  ex- 
amination will  show  to  be  entirely  amorphous  (distinction 
between  nicotine  and  coniine).  If  kept  for  a  long  time  in  a 
desiccator  over  sulphuric  acid,  it  will  become  indistinctly 
crystalline. 

2.  Roussin's  Test.- — Dissolve  a  trace  of  nicotine  in  ether, 
using  a  dry  test-tube.  Add  to  this  solution  about  the  same 
volume  of  ether  containing  iodine.  Stopper  and  set  the  test- 
tube  aside.  The  mixture  will  become  turbid  and  deposit  a 
brownish  red  resin  which  will  gradually  become  crystalline. 
After  some  time,  ruby-red  needles  with  a  dark  blue  reflex  will 
crystallize  from  the  ether.  These  are  "Roussin's  crystals." 
If  nicotine  is  old  or  resinous,  it  will  not  as  a  rule  give  these 
crystals. 

3.  Melzer's  Test.^ — If  a  drop  of  nicotine  is  heated  to  boiling 
with  2-3  cc.  of  epichlorohydrin,^  the  mixture  becomes  dis- 
tinctly red.     This  test  applied  to  coniine  causes  no  color. 

^  Zeitschrift  des  allgemeinen  Oesterreichen  Apotheker-Vereins  54,  65. 

CH2  CI  -  CH 
2  Epichlorohydrin,  |      /O,    prepared  by  the  action  of  i  mol.  of 

ch; 

caustic  alkali  on  a-dichlorohydrin,  CH2C1-CH(0H)-CH2C1,  or  a,  jS-dichloro- 
hydrin,  CH2(0H)-CHC1-CH2C1,  is  a  colorless  liquid  insoluble  in  water  and 
freely  soluble  in  alcohol  and  ether.  It  has  an  odor  like  chloroform  and  a  burning, 
sweetish  taste. 


NON-VOLATILE   POISONS  89 

4.  Schindelmeiser's  Test.^ — If  nicotine  that  is  not  resinous 
is  treated  first  with  a  drop  of  formaldehyde  solution  free  from 
formic  acid  and  then  with  a  drop  of  concentrated  sulphuric  acid, 
the  mixture  takes  on  an  intense  rose-red  color.  If  nicotine  and 
formaldehyde  are  in  contact  for  several  hours,  the  solid  residue 
obtained  gives  even  a  finer  color  reaction  with  a  drop  of  nitric 
acid.  Only  a  little  formaldehyde  should  be  used,  otherwise 
the  solution  becomes  green  after  a  while  and  decomposition 
takes  place. 

Under  the  same  conditions  trimethylamine,  piperidine,  pyridine,  picoline, 
quinoline  and  aniline  gave  no  color.  Nor  did  extracts  from  putrefying  horse-flesh 
and  the  entrails  of  animals,  poisoned  by  arsenic  or  mercury,  give  the  test,  at  least 
not  when  these  extracts  were  prepared  according  to  the  Stas-Otto  method. 

5.  Physiological  Test. — When  very  small  quantities  of  nico- 
tine are  present,  the  physiological  test  should  accompany  the 
chemical  tests.  A  very  characteristic  picture  is  given  by  frogs 
after  administration  of  small  doses  of  nicotine.  First  there  is 
stimulation,  then  paralysis  of  the  brain  and  respiratory  muscles 
and  apparent  curare-action  (tetanic  convulsions).  The  toxic 
action  of  pure  nicotine  should  be  studied  first.  The  experiment 
with  a  frog's  heart,  which  shows  temporary  cessation  of  diastole, 
is  also  very  characteristic. 

ANILINE 

AniHne,  C6H5.NH2,  upon  evaporation  of  the  ether  extract  from 
the  alkaline  solution,  will  usually  appear  as  reddish  or  brownish 
globules.  Dissolve  some  of  this  residue  in  water  and  apply 
the  aniHne  tests  already  described  on  page  45.  A  further 
test  for  aniline  consists  in  mixing  some  of  the  residue  with  a  few 
drops  of  concentrated  sulphuric  acid,  and  adding  a  few  drops 
of  potassium  dichromate  solution.  If  aniline  is  present,  an 
evanescent  blue  color  will  appear. 

VERATRINE 

Pure  officinal  veratrine  is  an  intimate  mixture  of  two  isomeric  alkaloids  ha\-ing 
the  composition  C32H49NO9.     These  are  cevadine,  also  called  cn,-stallized  vera- 

^  Pharmazeutische  Zentral-Halle  40,  703  (1S99). 


90  DETECTION   OF   POISONS 

trine,  which  is  nearly  insoluble  in  water;  and  amorphous  veratridine  which  is 
soluble  in  water.  Even  small  quantities  of  the  crystalline  alkaloid  will  render 
veratridine  insoluble  in  water.  On  the  other  hand  veratridine  will  prevent 
cevadine  from  crystallizing.  Consequently  the  crystalline  base  cannot  be  iso- 
lated by  recrystallizing  officinal  veratrine  from  alcohol  or  from  any  other  solvent; 
nor  can  the  water-soluble  alkaloid  be  obtained  by  simple  extraction  with  water. 

Separation  of  Cevadine  and  Veratridine. —E.  Schmidt  uses  the  following 
method  to  isolate  the  crystalline  and  the  water-soluble  veratrine  from  officinal 
veratrine.  Place  the  officinal  preparation  in  a  beaker  and  dissolve  in  strong 
alcohol.  Heat  this  solution  to  60-70°  and  add  enough  warm  water  to  produce  a 
permanent  turbidity.  Cautiously  add  just  enough  alcohol  to  clear  the  solution 
and  allow  evaporation  to  take  place  slowly  at  60-70°.  A  white,  crystalline  pre- 
cipitate will  presently  appear.  Filter  with  suction,  wash  the  precipitate  with  a 
little  dilute  alcohol  and  recrystaUize  from  hot  alcohol.  This  is  crystalline  vera- 
trine. Clear  the  filtrate  from  the  crystalline  precipitate  by  adding  a  little  alcohol 
and  evaporate  at  60-70°.  This  will  give  a  second  crop  of  crystals.  By  repeating 
this  process  several  times  one  may  obtain  in  a  crystalline  condition  about  one- 
third  of  the  veratrine  taken.  Finally  evaporate  the  filtrate  from  the  crystalline 
deposit  at  the  given  temperature  until  there  is  no  longer  any  odor  of  alcohol.  A 
considerable  quantity  of  a  resinous  mass  which  is  a  mixture  of  both  alkaloids  will 
separate.  The  aqueous  filtrate  from  this  deposit  will  contain  veratridine  which 
may  be  obtained  by  rapidly  evaporating  the  solution  in  vacuo  over  sulphuric  acid. 

Properties  of  Officinal  Veratrine. —Veratrine  appears  as  a  white,  amorphous 
powder  which  is  crystalline  under  the  microscope.  It  has  a  sharp,  burning  taste 
and  the  minutest  quantity  introduced  into  the  nostrils  excites  protracted  sneezing. 
It  is  almost  insoluble  in  boiling  water  and  the  aqueous  extract  always  has  a 
faintly  alkaline  reaction;  fairly  soluble  in  ether  (i  :  10),  benzene,  petroleum  ether 
and  amyl  alcohol;  and  freely  soluble  in  alcohol  (i  -.4)  and  chloroform  (i  :2). 
All  these  solutions  have  a  strong  alkaline  reaction.  Officinal  veratrine  melts  at 
150-155°  to  a  yellowish  Hquid  which  solidifies  to  a  transparent,  resinous  mass. 
If  the  veratrine  solution  is  faintly  acid,  ether  will  extract  a  very  little  of  the  alka- 
loid. Under  the  same  conditions,  chloroform  and  amyl  alcohol  will  extract 
more.  The  alkaloid  is  usually  deposited  from  ether  as  a  white,  amorphous  pow- 
der. Phospho-molybdic  acid,  iodo-potassium  iodide,  tannic  acid  and  potassium 
mercuric  iodide  give  distinct  precipitates  with  an  aqueous  veratrine  solution 
containing  hydrochloric  acid  and  diluted  i  :  5000.  Chlorides  of  gold  and  plati- 
num and  picric  acid  fail  to  show  the  alkaloid  in  this  dilution. 

Constitution. —Heated  with  saturated  barium  hydroxide,  or  alcoholic  potas- 
sium hydroxide  solution,  crystalHzed  veratrine  (cevadine)  is  hydrolyzed  into 
angelic  acid  and  cevine: 

C32H49NO9  +  H2O  =  C5H8O2I  +  C27H43NO8 

Cevadine  Angelic  Cevine 

acid 

^  Angelic  acid  (I)  and  tiglic  acid  (II)  are  stereo-isomers : 

I.  CH3— C— H  II.     H— C— CHsl 

CH3— C— COOH  CH3— C— COOH 


NON-VOLATILE    POISONS  91 

M.  Freund^  has  shown  that  ccvadinc  takes  up  only  one  acetyl  or  benzoyl 
group,  whereas  cevine  takes  up  two.  The  following  formula:  show  these 
relationships : 

/O.CsHtO  /OH 

C27H«N06<  '  -*  C27H4lN06< 

Cevadine  Cevine 

I  i 

/O.CsHyO  /O.CO.CH3 

C27H4iNOc<  CjyH^iNOe^ 

\O.CO.CH3  ^O.CO.CHs 

Acetyl-cevadine  Diacetyl-cevine 

By  means  of  hydrogen  peroxide  M.  Freund  has  converted  cevine  into  cevine 
oxide,  C27H43NO9,  which  crystallizes  well  and  contains  one  more  atom  of  oxygen. 

V 
This  compound  must  belong  to  the  class  of  the  amino-oxides,  i?  =  N  =  0,  for  sul- 
phurous acid  easily  converts  it  into  cevine. 

Detection  of  Veratrine 

1.  Concentrated  Sulphuric  Acid  Test. — Pour  a  few  drops  of 
concentrated  sulphuric  acid  upon  a  trace  of  veratrine.  The 
alkaloid  will  have  an  intense  yellow  color  and,  if  stirred,  will 
give  a  solution  of  the  same  color.  Gradually  this  color  will 
change  to  orange,  then  to  blood  red  and  finally  to  cherry  red. 
Gentle  heating  will  hasten  this  color  change  and  veratrine,  dis- 
solved in  concentrated  sulphuric  acid,  will  give  a  fine  cherry  red 
solution  almost  immediately. 

Frohde's  and  Erdmann's  reagents  give  color  changes  similar 
to  those  caused  by  sulphuric  acid. 

2.  Concentrated  Hydrochloric  Acid  Test. — If  a  trace  of  vera- 
trine is  dissolved  in  i  or  2  cc.  of  cold  concentrated  hydrochloric 
acid,  the  solution  will  be  colorless.  When  this  solution  is 
heated  10  to  15  minutes  in  a  boiling  water-bath,  a  cherry  red 
color  will  appear.  This  color  will  last  for  a  day  and  even  0.2 
mg.  of  veratrine  will  produce  it. 

3.  Concentrated  Nitric  Acid  Test. — Concentrated  nitric  acid 
dissolves  veratrine  with  a  yellow  color. 

4.  Weppen's  Test. — Thoroughly  mix  in  a  mortar  i  part  of 
veratrine  with  about  5  parts  of  finely  powdered  cane  sugar. 
Add  a  few  drops  of  concentrated  sulphuric  acid  to  some  of 

^Berichte  der  Deutschen  chemischen  Gesellschaft  37,  1946  (1904). 


92  DETECTION   OP   POISONS 

the  mixture  upon  a  watch  glass.  At  first  a  yellow  color  will 
appear  and  later,  beginning  at  the  margin  this  will  change  to 
grass  green  and  finally  to  blue.  Breathing  upon  the  mixture 
will  cause  the  color  to  change  more  quickly.  Too  great  an 
excess  of  cane  sugar  must  be  avoided. 

E.  Laves^  substitutes  an  aqueous  furfurol  solution  for  cane 
sugar  in  this  test.  Mix  in  a  test-tube  3  or  4  drops  of  i  per  cent, 
aqueous  furfurol  solution  with  i  cc.  of  concentrated  sulphuric 
acid.  Add  3  to  5  drops  of  this  solution  to  the  substance  to  be 
tested  so  that  it  just  touches  the  edge  of  the  liquid.  If  veratrine 
is  present,  a  dark  streak  will  gradually  run  from  the  substance 
into  the  liquid.  At  the  starting  point  it  will  appear  blue  or  blue 
violet  and  farther  away  green.  If  substance  and  liquid  are 
stirred  with  a  glass  rod,  the  liquid  will  become  dark  green. 
After  some  time,  or  more  quickly  when  warmed,  the  color 
will  become  blue  and  finally  violet. 

5.  Grandeau's  Test. — Direct  addition  of  1-2  drops  of  bro- 
mine water  to  the  yellow  solution  of  veratrine  in  concentrated 
sulphuric  acid  produces  an  immediate  purple  color  almost  iden- 
tical with  that  appearing  when  the  solution  of  the  alkaloid  in 
concentrated  sulphuric  acid  stands  a  long  time  or  is  gently 
warmed. 

6.  Vitali's  Test. — Dissolve  veratrine  in  a  few  drops  of  fuming 
nitric  acid  and  evaporate  the  solution  to  dryness  upon  the 
water-bath  in  a  porcelain  dish.  A  yellowish  residue  will  remain. 
If  this  is  cooled  and  then  moistened  with  an  alcoholic  potas- 
sium hydroxide  solution,  the  color  will  change  to  orange  red  or 
red  violet  and  stirring  will  produce  a  solution  having  the  same 
color. 

Atropine,  hyoscyamine,  scopolamine,  as  well  as  strychnine, 
respond  to  this  test  in  a  very  similar  manner, 

STRYCHNINE 

Strychnine,  C21H22N2O2,  occurs  with  brucine  chiefly  in  nux  vomica  and 
Ignatius  beans,  constituting  the  larger  part  of  the  mixed  alkaloids.  The  former 
contains  2.93-3.14  per  cent,  of  these  two  alkaloids  and  the  latter  3.11-3.22  per 

^  Pharmaceutische  Zeitung,  37,  338. 


NON-VOLATILE    POISONS  93 

cent.  The  free  base  strychnine  forn-js  colorless,  shining  prisms  belonging  to  the 
rhombic  system  which  melt  at  268°.  The  alkaloid  dissolves  in  6600  parts  of 
cold  and  2500  parts  of  hot  water,  giving  alkaline  solutions  having  a  very  bitter 
taste.  It  is  nearly  insoluble  in  absolute  alcohol  and  in  absolute  ether.  It  dis- 
solves in  160  parts  of  cold  and  1 2  parts  of  boiling  alcohol  (90  per  cent,  by  volume) ; 
it  is  also  soluble  in  commercial  ether  and  in  benzene;  but  most  readily  in  chloro- 
form (6  parts  at  15°).  Strychnine  diluted  with  water  i  :  Ooo.ooo  can  be  recognized 
by  its  bitter  taste. 

Strychnine  is  a  monacid  base  combining  with  one  equivalent  of  acid  and  form- 
ing salts  which  are  usually  crystalline.  These  salts  have  a  very  bitter  taste  and 
are  very  poisonous.  The  best  known  strychnine  salt,  and  one  used  medicinally, 
is  the  nitrate,  C21H22N2O2.HNO3.  The  combination  of  one  molecule  of  strych- 
nine with  one  molecule  of  an  alkyl  haloid,  for  example,  methyl  iodide,  to  form 
strychnine  iodo-methylate,  C21H22NO2.CH3.I,  shows  that  the  alkaloid  is  a  tertiary 
base.  Sodium  methylate  (CHs.ONa)  in  alcoholic  solution  converts  strychnine 
into  strychnic  acid  which  is  probably  an  imino-carboxylic  acid.  Strychnic  acid 
loses  a  molecule  of  water,  when  its  solutions  are  boiled  in  presence  of  mineral 
acids,  and  is  changed  to  strychnine.  Because  of  this  behavior  Tafel  regards 
strychnine  as  an  inner  anhydride  of  strychnic  acid,  one  containing  a  group  of  the 
character  of  an  acid  imide : 

(C2oH220)^CO  +  H2O  ^  (C2oH220)|-COOH 
\|  ^NH 

N 

Strychnine  Strychnic  acid 

On  the  basis  of  Tafel's  strychnine  formula,  strychnine  iodo-meth3date  would  be 
expressed  as  follows: 

/CH3 

#    ^I 
(C20H22O)— CO 

\l 

N 

Physiological  Action. — Strychnine  increases  reflex  irritability  of  the  brain  and 
spinal  cord.  Even  the  slightest  stimulus,  especially  if  acoustic,  optical,  or  tactile, 
may  cause  powerful  reflexes  after  large  doses  of  this  alkaloid.  Convulsions 
may  follow  each  stimulus,  if  the  dose  is  sufiicient.  Very  large  doses  of  strychnine 
cause  curare-like  paralysis  of  the  peripheral  ends  of  motor  nerves  in  frogs  and 
other  warm-blooded  animals.  It  may  also  affect  the  muscles  of  the  heart. 
Strychnine  diminishes  the  motile  power  of  leucocytes  and  then  arrests  their 
motion.  The  poison  also  affects  plant  protoplasm,  at  least  that  of  Mimosa 
pudica,  in  that  the  plant's  motor  organs  lose  their  elasticity  and  flexibility. 

Aside  from  the  saliva,  bile  and  milk,  the  urine  is  the  main  channel  through 
which  strychnine  is  eliminated  from  the  organism.  Human  urine  may  contain 
even  the  unaltered  alkaloid.  Elimination  begins  during  the  first  hour,  is  slight 
after  2  days  but  is  not  complete  until  much  later.  JMore  unaltered  str>'chnine  is 
eliminated  after  large  than  after  small  doses.  In  the  former  case  70-75  per  cent, 
of  the  alkaloid  may  remain  undecomposed.  The  liver,  kidneys,  brain  and  spinal 
cord  may  store  up  unchanged  strychnine.     (See  R.  Robert,  " Intoxikationen,") 


94  DETECTION   OF  POISONS 

Detection  of  Strychnine 

Sodium  and  potassium  hydroxide,  ammonia  and  alkaline 
carbonates  precipitate  the  free  base  strychnine  from  aqueous 
solutions  of  its  salts  as  a  white  crystalline  solid: 

C21H22N2O2.HNO3  +  NaOH  =  C21H22N2O2  +  H2O  +  NaNOs. 

Ether  will  extract  strychnine  from  an  alkaline  solution  and 
deposit  the  alkaloid  on  evaporation  in  fine  crystalline  needles. 
Chloroform  takes  up  the  alkaloid  more  freely,  since  strychnine 
is  considerably  more  soluble  in  this  solvent  than  in  ether.  Even 
very  dilute  solutions  of  strychnine  salts  give  precipitates  with 
most  of  the  alkaloidal  reagents.  Tannic  acid,  potassium  mer- 
curic iodide  and  phospho-tungstic  acid  produce  white  precipi- 
tates; gold  chloride  and  phospho-molybdic  acid  yellow  precipi- 
tates; and  iodo-potassium  iodide  brown  precipitates.  To 
obtain  tests  with  these  reagents,  the  residue  from  ether  should 
first  be  dissolved  in  very  dilute  hydrochloric  acid. 

Concentrated  sulphuric  acid,  Erdmann's  and  Frohde's 
reagents  dissolve  perfectly  pure,  brucine-free  strychnine  without 
color. 

Strychnine  is  soluble  in  concentrated  nitric  acid  with  a  yellow- 
ish color.  Potassium  dichromate,  added  to  solutions  of  strych- 
nine salts,  precipitates  strychnine  dichromate,  (C2iH22N202)2-- 
H2Cr207,  in  the  form  of  fine  yellow  crystalline  needles  which 
upon  recrystallization  from  hot  water  appear  as  shining  orange- 
yellow  needles. 

Potassium  ferricyanide,  added  to  solutions  of  strychnine 
salts,  precipitates  golden-yellow,  crystalline  strychnine  ferri- 
cyanide (C2lH22N202)2.H3Fe(CN)6  +  6H2O. 

Special  Reactions 

I.  Sulphuric  Acid-Dichromate  Test. — Dissolve  a  very  Kttle 
strychnine  in  2  or  3  drops  of  concentrated  sulphuric  acid 
upon  a  watch  glass.  The  solution  should  be  colorless.  Add  a 
fragment  of  potassium  dichromate  and  hold  it  firmly  in  one  place 
upon  the  glass.  Intense  blue  or  blue-violet  streaks  will  come 
from  the  potassium  dichromate,  if  the  watch  glass  is  tilted  up 


NON-VOLATILE   POISONS  95 

and  down.  If  the  entire  mixture  is  stirred,  the  sulphuric  acid 
will  have  a  beautiful  evanescent  blue  or  blue-violet  color. 

This  test  may  also  be  made  by  scattering  upon  the  surface  of 
the  solution  of  strychnine  in  concentrated  sulphuric  acid  a  few 
particles  of  coarsely  powdered  potassium  dichromate  and  mix- 
ing well  with  a  glass  rod.  In  this  way  the  blue  to  blue-violet 
color  reaction  is  given  very  beautifully.  The  blue  color  is  not 
permanent.     It  soon  changes  to  red  and  finally  to  dirty  green. ^ 

Strychnine  chromate  and  ferricyanide  give  this  test  especially 
well.  To  prepare  the  former  salt,  pour  a  very  dilute  potassium 
dichromate  solution  over  strychnine  upon  a  watch  glass.  When 
the  two  substances  have  been  in  contact  for  some  time,  pour 
the  remaining  liquid  from  the  strychnine  chromate.  Wash 
the  precipitate  once  with  a  little  water.  Put  some  strychnine 
chromate  upon  the  end  of  a  glass  rod  and  draw  it  through  a 
few  drops  of  concentrated  sulphuric  acid  upon  a  watch  glass. 
This  will  produce  violet  and  blue  streaks  in  the  acid. 

Mandelin's  reagent,^  that  is  to  say,  vanadic-sulphuric  acid, 
gives  this  strychnine  test  very  well.  The  blue  or  violet  color 
given  by  this  reagent  with  strychnine  is  more  permanent  than 
that  produced  by  potassium  dichromate.  The  color  finally 
changes  to  orange-red. 

Other  oxidizing  agents  may  be  substituted  for  potassium  dichromate,  as 
potassium  permanganate,  lead  peroxide,  manganese  dioxide,  potassium  ferri- 
cyanide (see  above),  cerium  oxide  and  vanadic  acid  (Mandelin's  reagent).  But 
neither  potassium  nitrate  nor  nitric  acid  can  be  used,  as  these  reagents  even  pre- 
vent this  test.     Consequently  strychnine  nitrate  does  not  give  the  test. 

2.  Physiological  Test. — Dissolve  the  ether  residue  in  a  little 
very  dilute  hydrochloric  acid.  Evaporate  the  filtered  solution 
to  dryness  upon  the  water-bath.  Dissolve  the  residue  in  pure 
water  (about  i  cc.)  and  inject  this  solution  into  the  lymph  sac 
on  the  back  of  a  lively  frog.  Keep  the  experimental  frog  in  a 
large,  loosely  covered  beaker.  Toxic  symptoms  will  appear  in 
5  to  30  minutes,  depending  upon  the  quantity  of  strychnine. 

1  According  to  Tafel  (Annalen  der  Chemie  und  Pharmazie,  268,  233  (iS92),this 
color  reaction  is  characteristic  of  manj^  anilides  and  is  due  to  the  presence  of  the 
group  —  CO— N  =  . 

2  See  page  314  for  the  preparation  of  this  reagent. 


96  DETECTION   OF   POISONS 

Strychnine  does  not  increase  reflex  irritability  for  all  kinds  of 
stimuli  but  only  for  tactile,  optical  and  especially  for  acoustic 
stimuli.  When  the  dose  of  strychnine  is  sufficiently  large, 
each  kind  of  stimulus  mentioned  will  produce  convulsions  like 
those  caused  by  tetanus.  For  example,  if  the  beaker  contain- 
ing the  "strychnine  frog"  is  gently  tapped,  this  slight  acoustic 
stimulus  is  sufficient  to  produce  convulsions. 

Detection  of  Strychnine  in  Presence  of  Brucine 

More  than  traces  of  brucine  prevent  detection  of  strychnine 
with  concentrated  sulphuric  acid  and  potassium  dichromate. 
Under  certain  conditions  Mandelin's  reagent  will  show  strych- 
nine more  or  less  distinctly  in  presence  of  brucine.  Dissolve 
the  ether  residue  in  concentrated  sulphuric  acid,  if  brucine  is 
present,  and  add  a  trace  of  concentrated  nitric  acid.  A  red 
color  indicates  brucine.  When  the  color  has  changed  to  yellow, 
add  a  fragment  of  potassium  dichromate  and  stir.  The  mixture 
will  become  blue  or  reddish  violet,  if  strychnine  is  present. 

Solid  potassium  permanganate,  stirred  witii  concentrated  sulphuric  acid  alone, 
will  give  a  dark  green  solution  which  is  yellowish  green  in  a  thin  layer  and  assumes 
with  time  a  red  to  violet  color  on  the  margin. 

The  same  procedure  used  to  estimate  these  two  alkaloids 
quantitatively  will  permit  detection  of  strychnine  even  in  pres- 
ence of  considerable  brucine.  Dissolve  the  residue  containing 
brucine  in  about  2  cc.  of  dilute  sulphuric  acid,  add  2  drops  of 
concentrated  nitric  acid  and  let  the  mixture  stand  4  hours. 
Render  alkaUne  with  excess  of  sodium  hydroxide  solution  and 
extract  thoroughly  with  ether.  The  residue  from  ether  will  be 
brucine-free  or  nearly  so.  Strychnine  thus  treated  will  give 
very  satisfactory  tests  with  concentrated  sulphuric  acid  and 
potassium  dichromate  and  with  Mandelin's  reagent. 

BRUCINE 

Brucine,  C23H26N2O4,  crystallizes  in  transparent,  monoclinic  prisms  or  shining 
leaflets.  Crystals  from  water  contain  either  4  or  2  molecules  and  from  alcohol 
2  molecules  of  water  of  hydration.  It  melts  in  its  water  of  hydration  only  a  few 
degrees  above  100°,  whereas  the  anhydrous  base  melts  at  178°.  Brucine  is 
more  readUy  soluble  than  strychnine  both  in  water  and  in  alcohol  and  therefore 


NON-VOLATILE    POISONS  97 

remains  dissolved  in  the  mother  liquors  from  the  preparation  of  strychnine.  It 
is  also  more  soluble  than  strychnine  in  ether.  Brucine  solutions  have  a  very 
bitter  taste  and  a  strong  alkaline  reaction.  Benzene,  but  especially  chloroform 
and  amyl  alcohol,  are  excellent  solvents  for  brucine.  Brucine  dilTers  from  strych- 
nine in  being  deposited  usually  amorphous  by  evaporation  of  its  ether  solution. 

Brucine  is  a  monacid,  tertiary  base  and  as  such  forms  addition  products  with 
one  molecule  of  an  alkyl  iodide.  For  example,  with  methyl  iodide  it  gives 
brucine  iodo-methylate,  C23H2cN04.N.Cri3l.  With  one  equivalent  of  acid 
brucine  gives  in  part  crystalline  salts.  Brucine  nitrate,  C23H2CN2O4.HNO3. 
2H2O,  crystallizes  in  rectangular  prisms. 

Brucine  may  be  shown  by  Zeisel's  method^  to  contain  two  methoxyl  groups 
(-OCH3). 

Heated  in  sealed  tube  to  80°  with  sodium  and  alcohol,  until  solution  is  complete, 

brucine  is  converted  into  brucic  acid,  C23H28N2O6.H2O,  which  contains  an  imino- 

group  ( =  NH)  in  its  molecule  since  it  forms  a  nitrosamine.     Tafel  and  Moufang^ 

express  the  relationship  between  brucine  and  brucic  acid  as  follows: 

N  N 

/-  # 

C2oH2o(OCH3)20— CO  +  H2O  ^  C2oH2o(OCH3)20— COOH 

\l  \ 

N  NH 

Brucine  Brucic  acid 

Heated  with  water,  brucic  acid  is  converted  into  brucine.  Consequently 
brucic  acid  is  related  to  brucine  as  strychnic  acid  is  to  strychnine. 

Detection  of  Brucine 
Ether,  benzene  or  chloroform  will  extract  brucine  from  an 
alkaline  solution.     Evaporation  of  the  ether  extract  usually 
leaves  the  alkaloid  in  an  amorphous  condition.     The  sensitive- 
ness of  the  alkaloidal  reagents  toward  brucine  is  as  follows : 

lodo-potassium  iodide  (i  :  50,000)  Potassium  bismuthous  iodide  (i  :  5000) 

Potassium  mercuric  iodide  (i  : 30,000)      Phospho-molybdic  acid  (i  :  2000) 
Gold  chloride  (i  :  20,000)  Tannic  acid  (i  :  2000) 

Platinic  chloride  (i  :  1000). 

1  Many  alkaloids  contain  one  or  more,  sometimes  three  or  more,  methoxyl 
groups  (—  OCH3)  united  with  a  benzene  nucleus.  The  determination  of  the 
number  of  such  groups  in  the  molecule  is  of  the  greatest  importance  as  a  step  in 
establishing  the  constitution  of  an  alkaloid,  because  in  this  way  some  of  the  car- 
bon, oxygen  and  hydrogen  atoms  are  at  once  disposed  of.  The  method  employed 
for  this  purpose  depends  on  the  fact  that  all  substances  containing  methoxjd 
groups  are  decomposed  by  hydriodic  acid,  yielding  methyl  iodide  and  a  hydroxyl 
compound.  By  estimating  the  methyl  iodide  obtained  from  a  given  quantity 
of  a  compound  of  known  molecular  weight,  it  is  easy  therefore  to  determine  the 
number  of  methoxyl  groups  in  the  molecule.  This  method  was  first  applied  by 
Zeisel  and  is  of  general  application.  (Perkin  and  Kipping,  "Organic  Chem- 
istry," page  498.) 

^  Annalen  der  Chemie  und  Pharmazie  304,  28  (1899). 
7 


98  DETECTION  OF  POISONS 

1.  Nitric  Acid-Stannous  Chloride  Test. — Concentrated  nitric 
acid  dissolves  brucine  and  its  salts  with  a  blood  red  color.  This 
color,  however,  is  slightly  stable  and  soon  changes  to  yellowish 
red  and  finally,  especially  with  heat,  to  yellow.  Add  a  few 
drops  of  freshly  prepared,  dilute  stannous  chloride  solution 
to  this  yellowish  red  or  yellow  solution.  An  intense  violet 
color  will  appear.  Heat  usually  changes  this  violet  color  again 
to  yellowish  red,  but  addition  of  a  few  more  drops  of  stannous 
chloride  solution  will  cause  the  violet  color  to  reappear.  The 
smaller  the  quantity  of  nitric  acid,  the  more  likelihood  that  this 
test  will  give  a  good  result.  Colorless  ammonium  sulphide 
solution  may  be  substituted  for  stannous  chloride. 

2.  R.  Mauch's  Modification  of  Nitric  Acid-Stannous  Chloride 
Test. — An  excellent  result  can  be  obtained  with  this  test  in  the 
following  manner.  Dissolve  brucine  in  60  per  cent,  aqueous 
chloral  hydrate  solution  and  put  about  0.5  cc.  of  this  solution 
into  a  test-tube.  Add  very  little  dilute  nitric  acid  and  thor- 
oughly mix  the  two  solutions.  Add  this  mixture  to  3  times 
its  volume  of  concentrated  sulphuric  acid  so  that  the  former  is 
on  the  surface.  A  yellowish  red  to  deep  red  zone,  depending 
upon  the  quantity  of  brucine,  will  appear  immediately.  When 
the  upper  layer  becomes  yellow,  introduce  by  a  pipette  a  little 
stannous  chloride  solution^  as  a  top  layer.  A  brilliant,  intensely 
violet  zone  will  appear  between  the  two  upper  layers.  The 
intensity  of  this  color  will  gradually  increase,  especially  if  the 
test-tube  is  gently  tilted  to  and  fro. 

ATROPINE 

H2C— CH CH2  CH2OH 

I  I  I 

N.CH3   CH— O.CO— CH 

I  I  I 

H2C— CH CH2  CfiHs 

Atropine,  C17H23NO3,  crystallizes  in  shining  pointed  needles  which  melt  at  115° 
and  dissolve  in  600  parts  of  water,  50  parts  of  ether  and  3.5  parts  of  chloroform. 
It  is  also  soluble  in  alcohol,  amyl  alcohol  and  benzene.     The  aqueous  solution  of 

^  Prepare  stannous  chloride  solution  by  dissolving  i  part  of  stannous  chloride 
in  9  parts  of  hydrochloric  acid  having  a  specific  gravity  of  1.12  (about  24  per 
cent.  HCl). 


NON-VOLATILE   POISONS 


99 


the  alkaloid  is  alkaline  and  has  a  lasting,  unpleasant,  bitter  taste.     Unlike  the 
optically  active  hyoscyaminc,  atropine  is  inactive. 

Constitution. — Heated  with  hydrochloric  acid  at  120-130°,  atropine  is  decom- 
posed into  tropic  acid  and  tropine: 


H 


H2C- 


H 

-C- 


-CH, 


H3C— N     HC— O 


H2C- 


H 


-CH 

Atropine 


OH 


H 
-C- 


-CH2 


CH2— OH    H2C- 

C— CH  =        H3C— N     HC— OH  + 

II      I                         III 
O    CeHs  H2C C CH2 


C 

H 

Tropine 


•   '  CH2— OH 

I 
HO— C— CH 

II      I 

O     CeHs 
Tropic    acid 

Heated  with  barium  hydroxide  solution,  atropine  yields  atropic  acid  which  is 
unsaturated  and  differs  from  tropic  acid  by  one  molecule  of  water: 


CH2.0H 

1 

CH2 

II 

CH.CcHb  - 

H2O 

ll 
=  CCeHs 

COOH 

Tropic  acid 

COOH 

Atropic  acid 

Since  the  structure  both  of  tropine  and  tropic  acid  has  been  determined  by 
synthesis  as  well  as  by  decomposition,  that  of  atropine  is  also  known.  Nitro- 
gen in  atropine  is  in  the  tertiary  condition.  Hyoscyamine  is  the  stereo-isomer  of 
atropine.  The  former,  heated  at  110°  out  of  contact  with  air,  or  allowed  merely 
to  stand  in  alcoholic  solution  with  addition  of  a  few  drops  of  an  alkaline  hydroxide 
solution,  is  changed  to  inactive  atropine.  Atropine  most  likely  is  the  racemic 
form,  whereas  hyoscyamine  is  the  laevo-rotatory  modification  of  this  isomeric 
base.  The  degree  of  rotation  of  hyosc3'amine  is  [a.]j)  =  —20.97°.  Toward 
alkaloidal  reagents  and  when  heated  with  concentrated  sulphuric  acid,  hy- 
oscyamine behaves  like  atropine.  It  also  resembles  the  latter  in  gi\dng  VitaU's 
reaction  (see  below). 

Putrefaction. — Ipsen^  has  found  atropine  very  resistant  in  presence  of  putre- 
fying material.  Even  after  2  years  he  could  detect  the  alkaloid  which  had  been 
exposed  to  the  influences  of  decomposition.  He  experimented  vrith  0.05  gram 
of  atropine  sulphate  in  respectivelj'^  300  cc.  of  blood,  urine  and  beer  and  with  pure 
atropine  in  300  cc.  of  blood. 

^  Vierteljahrsschrift  fiir  gerichtliche  Medizin  und  offentliches  Sanitatswesen^ 
31,  308. 


100  DETECTION    OF   POISONS 

Detection  of  Atropine 

Ether,  benzene  or  chloroform  will  extract  atropine  from  a 
solution  alkaline  with  sodium  hydroxide  or  carbonate  solution. 
In  a  special  search  for  atropine  use  sodium  carbonate  solution 
and  extract  with  chloroform  which  is  a  better  solvent  than  ether. 
Evaporate  the  solvent  and  test  the  residue,  which  is  usually 
amorphous,  as  follows: 

1.  Vitali's  Test. — Dissolve  the  alkaloid  in  a  few  drops  of 
fuming  nitric  acid,  and  evaporate  the  solution  in  a  porcelain 
dish  to  dryness  upon  the  water-bath.  Moisten  the  yellowish 
residue  when  cold  with  a  few  drops  of  a  solution  of  potassium 
hydroxide  in  absolute  alcohol.  An  evanescent  violet  color 
will  appear,  if  atropine  is  present. 

Hyoscyamine  and  scopolamine  also  give  Vitali's  test.  Strychnine  and  vera- 
trine  behave  similarly.  This  test  therefore  is  characteristic  of  the  atropine 
alkaloids  only  in  the  absence  of  the  two  latter  alkaloids. 

2.  Odor  Test. — Heat  a  little  atropine  in  a  dry  test-tube  until 
a  white  vapor  appears.  An  agreeable  odor  will  arise  at  the  same 
time.  Then  add  about  i  cc.  of  concentrated  sulphuric  acid,  and 
heat  until  the  acid  begins  to  darken.  Dilute  at  once  with 
about  2  cc.  of  water.  During  the  foaming  there  will  be  an  in- 
tense, sweetish  odor  like  that  of  honey.  By  this  test,  which 
was  formerly  the  only  method  of  identifying  atropine,  o.oi 
gram  of  the  alkaloid  can  be  detected. 

3.  Physiological  Test. — Atropine  acts  in  a  very  characteristic 
manner  upon  the  pupil  of  the  eye,  and  this  behavior  can  be 
-employed  as  a  test.  One  drop  of  an  atropine  solution  diluted 
1 :  130,000  will  produce  a  noticeable  enlargement  of  the  pupil. 
Dissolve  a  small  portion  of  the  ether  residue  in  4  or  5  drops  of 
very  dilute  sulphuric  acid,  and  introduce  a  drop  of  this  solution 
into  a  dog's  or  a  cat's  eye.  The  enlargement  of  the  pupil  often 
persists  for  several  hours.  The  utmost  care  should  be  taken 
in  performing  this  test,  if  applied  to  the  human  eye. 

The  following  alkaloidal  reagents  are  especially  sensitive 
toward  atropine:  iodo-potassium  iodide,  phospho-molybdic 
acid   (1:10,000),  gold  chloride,  phospho-tungstic  acid,  potas- 


NON-VOLATILE   POISONS  101 

sium  mercuric  iodide,  potassium  bismuthous  iodide.  Picric 
acid,  added  to  solutions  of  atropine  salts  that  are  not  too 
dilute,  will  precipitate  atropine  picrate  as  yellow  leaflets. 
Platinic  chloride  gives  monoclinic  prisms. 

HOMATROPINE 

Homatropine,  C16H21NO3,  is  the  tropyl  ester  of  phenyl-glycolic  or  mandelic 
acid.     The  hydrochloride  of  this  base  is  obtained  by  heating  a  mixture  of  tropine, 

TT  p pjT         pxT  p  XT      mandelic  acid  and  hydrochloric  acid,  the  latter 

I  I  I  acting  as  a  dehydrating  agent. 

N.CH3   CH.OOC— CH  The  hydrobromide  of  homatropine  (CieH2i- 

I  I  '  NOs.HBr)  is  used  in  medicine  as  a  substitute 

^  2  for  atropine.     Its  action  on  the  pupil  is  nearly 

as  strong  as  that  of  the  natural  alkaloid  and  its  effect  disappears  in  12-24  hours, 
whereas  that  of  atropine  often  lasts  8  days.  Moreover  it  is  less  toxic  than 
atropine.  Homatropine  is  a  strong  tertiary  base  which  forms  neutral  salts  with 
acids.  This  alkaloid  does  not  give  Vitali's  test.  It  melts  at  92-96°;  hj-oscya- 
mine  at  108°;  and  atropine  at  115.5°. 

OfHcinal  homatropine  hydrobromide  may  be  distinguished  from  the  hydro- 
bromides  of  atropine  and  hyoscyamine  by  warming  the  substance  in  a  test-tube 
with  a  little  chloroform.  This  solvent  dissolves  the  latter  two  salts  in  every  pro- 
portion but  homatropine  hydrobromide  is  insoluble.  An  alternative  procedure 
consists  in  dissolving  the  given  salt  in  a  little  water,  precipitating  the  base  with 
sodium  carbonate  solution,  and  extracting  with  ether.  Dehj^drate  the  ether 
extract  with  potassium  carbonate  and  evaporate  slowly  in  a  moderately  warm 
place.  This  method  will  give  crystals  of  the  alkaloid.  Dry  these  crystals  in 
vacuo  over  concentrated  sulphuric  acid  and  determine  their  melting  point. 

COCAINE 

Cocaine,  Ci7H2iN04,  crystallizes  from  alcohol  in  large,  colorless,  monoclinic 
prisms  which  melt  at  98°.     It  has  a  bitterish  taste  and,  placed  upon  the  tongue, 

XT  p pTT QTT rooCH  causes    temporary,    local    anaesthesia.      The 

I  [  alkaloid   is    only    slightly   soluble    in    water 

N.CH3    CH— OOC— CeHs    (1:700),  but  easily  soluble  in  alcohol,  ether, 
TT  „      '  TT         /Itt  chloroform,  benzene  and  acetic  ether.      Its 

^  ^  solutions  are  strongly  alkaline  and  Isevo-rota- 

tory.  Dilute  acids  easily  dissolve  cocaine  and  in  most  cases  form  readily  cr>'S- 
tallizable  salts.  The  fixed  alkalies,  ammonia  and  alkaline  carbonates  precipitate 
the  free  base  from  solutions  of  its  salts. 

Constitution. — Cocaine  is  a  monacid,  tertiarj^  base,  since  it  adds  a  molecule  of 
CH3I.  On  distillation  with  barium  hydroxide,  this  alkaloid  loses  methyl  amine 
(CH3.NH2),  thus  proving  the  attachment  of  a  method  group  to  nitrogen.  Co- 
caine must  therefore  contain  the  group  =  N — CH3.  This  base  is  also  the 
methyl  ester  of  an  acid  and  at  the  same  time  the  benzojd  derivative  of  an  alcohol, 
for  it  is  decomposed  into  benzoyl-ecgonine  and  methjd  alcohol  when  heated  with 


102 


DETECTION   OF   POISONS 


water.  If  mineral  acids,  barium  hydroxide  or  alkalies  are  used  instead  of  water, 
the  primary  product,  benzoyl-ecgonine,  is  further  decomposed  into  ecgonine, 
benzoic  acid  and  methyl  alcohol.  Taking  the  structural  formula  proposed  by 
Willstatter,  we  may  express  this  reaction  as  follows: 

Cocaine 
H2C CH2 


CH3 


HC— N- 

I      H 


-CH 


H2C— C— CH 


Ecgonine 

Benzoic 
acid 

Methyl 
alcohol 

H2C CH2 

CeHs 

CH3 

CH3 

1 

1 

1 

CO 

0 

HC— N— CH 

1 

H 

H 

0 
H 

H2C— C— CH 

1        1 

+ 

+ 

1        1 
0     CO 

1         1 

1       1 
H      0 

H 

CeHs— CO  -0     CO— OCHs 
+ 
HOiH       HO     :H 

Water 
Ecgonine  (I)  heated  with  phosphorus  oxychloride  loses  a  molecule  of  water  and 
passes  into  anhydro-ecgonine  (II).  The  latter  heated  to  280°  with  fuming 
hydrochloric  acid  loses  carbon  dioxide  and  is  converted  into  tropidine  (III). 
Tropidine  heated  with  a  caustic  alkaUne  solution  adds  a  molecule  of  water  and 
passes  into  tropine  (IV) ,  the  basic  cleavage  product  of  atropine.  Evaporation  of 
tropine  with  tropic  acid  in  dilute  hydrochloric  acid  solution  yields  atropine  (V). 
Thus  it  is  possible  to  start  with  the  alkaloid  cocaine  and  synthesize  the  alkaloid 
atropine.  The  series  of  changes  involved  is  as  follows : 
H2C— CH CH.COOH  H2C— CH CH.  iCOO;  H 


N.CH3  CH. 


OH  i   -  H2O 
H     i 


H2C— CH CH 

Ecgonine  (I) 

H2C — CH CH2 

N.CH3  CH     +  H2O 


N.CH3  CH 

il 


-  CO2 


H2C— CH CH 

Anhydro-ecgonine  (II) 

H2C— CH CH2  CH2.OH 

i  I  ., - -:  I 

N.CH3  CH.O:  H  +  HO  iOC-CH 


H2C— CH CH 

Tropidine  (III) 


H2C— CH CH 


H2C — CH CH2 

Tropine  (IV) 

CH2.OH 


CeHfi 

Tropic  acid 


N.CH3  CH.O.CO.CH 


CfiHs 


+  H2O. 


H2C — CH CH2 

Atropine  (V) 
Behavior  in  the  Animal  Organism. — Experiments  upon  dogs  and  rabbits  show 
that  the  former  animal  eliminates  through  the  kidneys  not  more  than  5  per  cent, 
of  the  cocaine  as  such  and  the  latter  none  at  aU.  As  the  urine  of  these  animals 
also  contains  no  ecgonine,  the  supposition  is  that  the  alkaloid  is  profoundly 
changed  in  the  animal  organism.  The  same  is  true  of  the  human  organism. 
Proells^  was  able  to  detect  cocaine  in  cadaveric  material  at  most  after  14  days. 
In  the  living  organism  the  alkaloid  is  said  to  be  changed  rapidly  into  ecgonine. 

1  Apotheker-Zeitung  16,  779,  788. 


NON-VOLATILE   POISONS  103 

Detection  of  Cocaine 

Ether,  chloroform  or  benzene  will  extract  cocaine  from  an 
alkaline  aqueous  solution.  Most  of  the  alkaloidal  reagents  will 
precipitate  cocaine  even  from  very  dilute  solutions  of  its  salts. 
The  reagents  especially  sensitive  are:  iodo-potassium  iodide, 
phospho-molybdic  and  phospho-tungstic  acids,  potassium  mer- 
curic iodide,  potassium  bismuthous  iodide,  gold  and  platinum 
chlorides,  and  picric  acid. 

Pure  concentrated  sulphuric  and  nitric  acids,  as  well  as  Erd- 
mann's,  Froehde's  and  Mandelin's  reagents,  dissolve  cocaine 
without  color. 

1.  Precipitation  Test. — If  i  or  2  drops  of  potassium  hydroxide 
solution  are  added  to  an  aqueous  solution  of  a  cocaine  salt  not 
too  dilute,  it  will  become  milky.  First,  resinous  globules  and 
later  fine,  crystalline  needles  of  the  free  base,  cocaine  (melting 
point  98°),  separate  from  solution: 

Ci7H2iN04.HCli  +  KOH  =  C17H21NO4  +  H2O  +  KCl. 

In  applying  this  test  to  the  ether  residue,  dissolve  a  consider- 
able quantity  in  a  few  drops  of  dilute  hydrochloric  acid  and  add 
potassium  hydroxide  solution  drop  by  drop  until  alkaHne  and 
cool  well  by  setting  in  ice.  Special  care  must  be  taken  to  have 
the  alkaloid  pure  enough  when  dry  for  a  melting-point  deter- 
mination. 

This  test  is  not  characteristic  of  cocaine  (except  the  melting 
point  which,  however,  requires  considerable  pure  material), 
because  most  of  the  alkaloids  are  precipitated  by  potassium 
hydroxide  solution  in  much  the  same  way. 

2.  Potassium  Permanganate  Test. — Add  saturated  potassium 
permanganate  solution  drop  by  drop  to  a  concentrated  aqueous 
solution  of  a  cocaine  salt.  This  reagent  will  give  a  violet, 
crystalline  precipitate  of  cocaine  permanganate.  In  applpng 
this  test  to  the  ether  residue,  dissolve  a  considerable  quantity 
in  2  drops  of  dilute  hydrochloric  acid  and  evaporate  the  solution 

^  Cocaine  hydrochloride  crystallizes  from  a  concentrated  aqueous  solution  in 
fine  prisms  containing  2  molecules  of  water  which  are  easily  given  off.  This  salt 
crystallized  from  alcohol  is  anhydrous  and  has  the  formula  C17H21NO4.HCI. 
The  anhydrous  compound  is  the  officinal  salt. 


104  DETECTION   OF   POISONS 

upon  the  water-bath.     Dissolve  the  residue  in  as  little  water 
as  possible  and  add  potassium  permanganate  solution. 

3.  Chromic  Acid  Test. — Add  a  few  drops  of  a  5  per  cent, 
chromic  acid  solution,  or  potassium  dichromate  solution  of 
corresponding  concentration  (7.5  per  cent.)  to  a  solution  of  a 
cocaine  salt.  Each  drop  will  produce  a  precipitate  which  will 
immediately  disappear  if  the  solution  is  shaken.  Then  add 
to  the  clear  solution  about  i  cc.  of  concentrated  hydrochloric 
acid  which  will  produce  an  orange-yellow  precipitate  more  or 
less  crystalline. 

4.  Detection  of  Benzoyl  Group. — This  test  requires  at  least 
0.2  gram  of  cocaine.  First,  digest  the  cocaine  a  few  minutes 
in  a  test-tube  with  2  cc.  of  concentrated  sulphuric  acid  upon  a 
boiling  water -bath.  Cool  and  dilute  with  a  little  water,  all 
the  while  keeping  the  mixture  cold.  A  white  crystalline  pre- 
cipitate of  benzoic  acid  will  appear.  Collect  and  dry  this 
precipitate  upon  a  filter.  Benzoic  acid  may  be  recognized  by 
subhming  the  precipitate,  or,  if  the  quantity  is  sufiicient,  by 
determining  the  melting  point  (120°). 

Benzoic  acid  may  also  be  extracted  with  ether.  Mix  the 
residue,  obtained  by  evaporating  the  solvent,  with  i  cc.  of 
absolute  alcohol  and  the  same  quantity  of  concentrated  sul- 
phuric acid.  The  characteristic  odor  of  ethyl  benzoate, 
C6H5.CO.OC2H5,  will  be  recognized. 

5.  Reichard's^  Test. — Addition  of  a  concentrated  aqueous 
solution  of  sodium  nitroprusside,  Na2Fe(CN)6N0.2H20,  drop 
by  drop  to  a  cocaine  salt  solution,  containing  at  least  4  mg. 
of  cocaine  per  cc,  causes  an  immediate  turbidity  which  will 
appear  under  the  microscope  as  well  formed  reddish  crys- 
tals. These  crystals  will  dissolve,  if  the  liquid  is  warmed,  and 
appear  again  if  the  solution  is  well  cooled.  Morphine  does  not 
give  this  test. 

6.  Goeldner's^  Test. — Mix  about  0.005  gram  of  pure  resor- 

^  C.  Reichard,  Chemiker-Zeitung  28,  299  (1904).  Pharmazeutische  Zeitung 
1904,  Nr.  29.     Pharmazeutische  Zentralhalle  45,  645  (1904). 

^  Pharmazeutische  Zeitschrift  fiir  Russland  28,  489  and  Zeitschrift  fiir  analy- 
tische  Chemie  40,  820  (1901). 


NON-VOLATILK   POISONS  105 

cinol  (C6H4(OH)2  1,3)  in  a  small  porcelain  dish  with  5-6  drops 
of  pure  concentrated  sulphuric  acid.  Add  about  0.02  gram  of 
cocaine  hydrochloride  to  this  solution  which  usually  has  a  faint 
yellowish  color.  There  is  a  vigorous  reaction,  during  which  the 
liquid  acquires  a  fine  blue  color  like  that  of  the  corn  flower. 
The  intensity  of  this  color  gradually  increases.  Sodium  hydrox- 
ide solution  will  change  the  blue  to  light  pink. 

7.  Physiological  Test. — Dissolve  the  material  (the  residue 
from  the  ether  extract)  in  a  few  drops  of  dilute  hydrochloric 
acid  and  evaporate  the  solution  to  dryness  upon  the  water-bath. 
Dissolve  the  residue  in  a  little  pure  water  and  apply  this  solu- 
tion to  the  tongue.     Cocaine  produces  a^  temporary  anesthesia. 

R.  Kobert  ("  Intoxikationen ")  has  found  small  frogs  suffi- 
ciently sensitive  for  use  in  the  physiological  test  for  cocaine. 
The  effects  to  be  observed  are  dilatation  and  fixedness  of  the 
pupil,  enlargement  of  the  palpebral  fissure  and  also  stimulation 
of  the  nervous  system.  Administer  the  same  quantity  of  co- 
caine hydrochloride  to  animals  for  comparison. 

PHYSOSTIGMINE 

Physostigmine,  C16H21N3O2,  also  called  eserine,  occurs  in  the  Calabar  bean, 
the  seed  of  Physostigma  venenosum.  This  alkaloid  is  deposited  from  benzene 
solution  upon  spontaneous  evaporation  of  the  solvent  in  large,  apparently 
rhombic  crystals  melting  at  105°.  Though  but  slightly  soluble  in  water,  it 
dissolves  freely  in  alcohol,  ether,  benzene  or  chloroform.  Physostigmine  solu- 
tions are  strongly  alkahne,  almost  tasteless  and  Isevo-rotatory.  It  is  a  strong 
monacid  tertiary  base,  forming  salts  with  acids  that  easily  undergo  decomposition 
and  crystallize  with  difficulty.  Light  and  heat  cause  acid  and  alkaline  solutions 
of  this  alkaloid  to  turn  red.  Owing  to  this  tendency  of  phj'sostigmine  to  undergo 
decomposition,  care  must  be  taken  during  its  isolation  to  keep  it  from  light  and  air 
and  also  to  avoid  rise  of  temperature.  Exclusion  as  far  as  possible  of  free  min- 
eral acids  and  caustic  alkalies  is  also  desirable. 

Detection  of  Physostigmine 

Concentrated  sulphuric  and  nitric  acids  dissolve  physostigmine  with  a  j'eUow 
color  which  soon  changes  to  olive-green.  The  alkaloid  evaporated  upon  the 
water- bath  with  fuming  nitric  acid  leaves  a  residue  ha^-ing  a  green  margin. 
Water,  alcohol  and  sulphuric  acid  dissolve  this  residue  with  a  green  color. 

1.  Ammonia  Test. — If  a  small  quantity  of  a  phj-sostigmine  salt  is  evaporated 
to  dryness  upon  the  water-bath  with  ammonium  hj'droxide  solution,  a  blue  or 
blue-green  residue  will  remain.     This  wiU.  dissolve  in  alcohol  ^-ith  a  blue  color. 


106  DETECTION    OF   POISONS 

Excess  ot  dilute  mineral  acid,  or  acetic  acid,  added  to  this  solution  will  change  the 
color  to  red.  The  solution  is  also  strongly  fluorescent.  Examined  spectro- 
scopically  the  blue  alkaline  solution  shows  one  absorption  band  in  the  red;  and 
the  red  acid  solution  one  absorption  band  in  the  yellow. 

A  drop  of  concentrated  sulphuric  acid,  added  to  the  blue  residue  frOm  evapora- 
tion with  ammonia,  will  give  a  green  solution.  The  green  color  diluted  with 
alcohol  will  change  to  red.  If  the  alcohol  is  evaporated,  the  green  color  will 
reappear. 

2.  Rubreserine  Test. — If  an  aqueous  solution  of  a  physostigmine  salt  is  shaken 
for  some  time  with  an  excess  of  potassium  or  sodium  hydroxide  solution,  a  red 
coloring  matter,  rubreserine  (C13H16N2O2),  is  formed.  This  compound  separates 
as  red  needles  which  become  greenish  blue  on  further  oxidation  owing  to  formation 
of  eserine  blue. 

Barium  hydroxide  solution  may  be  substituted  for  the  caustic  alkali.  This 
reagent  first  produces  a  white  precipitate  which  soon  becomes  red  on  being  shaken. 
Sometimes  this  change  occurs  even  in  the  cold  but  invariably  takes  place  with 
heat. 

3.  Physiological  Test. — The  marked  action  of  physostigmine  in  causing  con- 
traction of  the  pupil  is  very  characteristic.  It  is  advisable  to  use  the  cat's  eye 
for  this  test.     Even  o.  i  mg.  of  this  alkaloid  wiU  produce  noticeable  contraction. 

CODEINE 

Codeine,  Ci7Hi8(CH3)N03,  the  methyl  ether  of  morphine,  crystallizes  from 
water,  or  from  ether  containing    water,  in  colorless,  transparent  octahedrons 
(--jj  which  are  often  very  large.     These  crystals  are  quite 

easily  soluble  in  water.  One  part  of  the  free  base 
is  soluble  at  15°  in  80  parts  of  water  and  at  100°  in 
15  parts.  Codeine  differs  from  most  of  the  other 
alkaloids,  morphine,  for  example,  in  its  relatively 
high  solubility  in  water.  Alcohol,  ether,  amyl  alcohol, 
chloroform  and  benzene  also  dissolve  codeine  freely. 
It  is,  however,  practically  insoluble  in  petroleum 
ether.  Aqueous  codeine  solutions  are  strongly  alka- 
line and  bitter.  Pure  codeine  does  not  reduce  iodic 
acid,  nor  does  it  immediately  produce  a  blue  color  or 
a  blue  precipitate  in  a  mixture  of  potassium  ferri- 
cyanide  and  ferric  chloride  solutions.  A  pure 
not  colored  blue  by  ferric  chloride  solution  alone. 
(Difference  between  morphine  and  codeine.)  Phospho-molybdic  acid,  iodo- 
potassium  iodide,  potassium  bismuthous  iodide  and  potassium  mercuric  iodide 
give  precipitates  even  with  very  dilute  codeine  solutions.  On  the  other  hand 
tannic  and  picric  acids,  gold  and  platinum  chlorides  are  less  sensitive. 

Detection  of  Codeine 

I.  Siilphuric  Acid  Test. — Concentrated  sulphuric  acid  dis- 
solves codeine  without  color.     After  long  contact  or  upon  appli- 


H     H2     1 
C      C      N 

HC      C      C      CH2 

CH3O.C     C      C     CH2 
C      C      CH 

0-HC     CH2 

\/ 

c 

/\ 

H     OH 

codeine   solution  is  also 

NON-VOLATILE   POISONS  107 

cation  of  gentle  heat,  the  solution  will  have  a  reddish  to  bluish 
violet  color.  The  solution  of  codeine  in  concentrated  sulphuric 
acid,  heated  to  about  150°  and  then  cooled,  is  colored  deep  red 
by  a  drop  of  concentrated  nitric  acid. 

2.  Nitric  Acid  Test. — Cold  nitric  acid  (25  per  cent.)  will 
convert  codeine  into  nitro-codeine  (Ci8H2o(N02)N03).  At  the 
same  time  the  acid  will  dissolve  the  alkaloid  with  a  yellow  color 
which  soon  changes  to  red.  Concentrated  nitric  acid  dissolves 
codeine  with  a  reddish  brown  color. 

3.  Oxidation  Test. — Mix  a  Httle  codeine  upon  a  watch  glass 
with  four  times  the  quantity  of  finely  powdered  potassium  arsen- 
ate (KH2ASO4).  Add  a  few  drops  of  concentrated  sulphuric 
acid  and  then  warm  gently  over  a  small  flame.  The  acid  will 
have  a  deep  blue  or  blue- violet  color,  if  the  codeine  is  not  quite 
pure.  Excess  of  potassium  arsenate  does  not  affect  the  test. 
If  water  or  sodium  hydroxide  solution  is  added,  the  blue  color 
will  change  to  orange-yellow. 

A  trace  of  ferric  chloride  solution  may  be  substituted  for  potassium  arsenate. 
Sulphuric  acid  containing  i  drop  of  ferric  chloride  solution  to  lo  cc.  of  acid  is 
prescribed  by  the  German  Pharmacopoeia  for  detecting  the  alkaloid  in  codeine 
phosphate. 

4.  Froehde's  Test. — This  reagent  dissolves  codeine  with  a 
yellowish  color  which  soon  changes  to  green  and  finally  to  blue. 
Gentle  warming  of  the  solution  over  a  very  small  flame  will 
hasten  this  change  of  color. 

R.  Mauch  warms  2  or  3  drops  of  a  chloral  hydrate  solution  of  codeine  with  i 
drop  of  Froehde's  reagent.     An  intense  blue  color  finally  appears. 

5.  Formalin-Sulphuric  Acid  Test.^ — Concentrated  sulphuric 
acid  containing  formalin  dissolves  codeine  with  a  reddish  violet 
color  which  changes  to  blue-violet.  This  color  is  persistent. 
The  spectrum  shows  an  absorption  of  orange  and  yellow. 

6.  Furfurol  Test.^ — Dissolve  codeine  in  a  few  drops  of  con- 

^  See  preparation  of  reagents,  page  314. 

^  This  test  depends  upon  furfurol  formed  by  the  action  of  concentrated  sul- 
phuric acid  upon  cane  sugar.  Very  dilute  aqueous  furfurol  solution  (i  :  1000) 
may  be  substituted  for  cane-sugar.  Excess  of  furfurol  unlike  cane-sugar  does 
not  interfere  vnth.  the  test.     Tr. 


108  DETECTION   OF   POISONS 

centrated  sulphuric  acid  and  warm  very  gently  with  a  drop  of 
cane-sugar  solution  which  must  not  be  in  excess.  This  will 
produce  a  purple-red  color. 

This  test  may  also  be  made  by  mixing  a  drop  of  sugar  solution 
with  codeine,  dissolved  in  about  5  drops  of  50-60  per  cent,  aque- 
ous chloral  hydrate  solution,  and  then  adding  1-2  cc.  of  con- 
centrated sulphuric  acid  as  an  underlayer.  A  carmine  red 
ring  will  appear  at  the  zone  of  contact.  The  color  is  quite 
permanent  and  increases  in  intensity  upon  standing.  If  the 
sulphuric  acid  and  chloral  hydrate  solution  are  thoroughly 
mixed,  the  entire  liquid  will  be  red.  After  a  time  the  shade  of 
color  will  be  more  of  a  red-brown. 

7.  Pellagri's  Test. — Both  codeine  and  morphine  give  this 
test.  Dissolve  codeine  in  concentrated  hydrochloric  acid  and 
add  at  the  same  time  3-4  drops  of  concentrated  sulphuric  acid. 
Expel  hydrochloric  acid  upon  the  water-bath  and  heat  the  resi- 
due about  15  minutes.  Dissolve  the  dirty  red  or  violet  residue 
in  2-3  cc.  of  water,  add  a  few  drops  of  hydrochloric  acid  and 
neutralize  with  acid  sodium  carbonate.  Then  add  alcoholic 
solution  of  iodine  drop  by  drop  (2  to  4  drops)  and  shake  thor- 
oughly for  several  minutes.  An  emerald  green  solution  indi- 
cates codeine.  Extract  the  green  solution  with  ether.  The 
color  of  the  ether  will  be  red,  whereas  that  of  the  aqueous  solu- 
tion will  remain  green.  This  is  a  test  for  apomorphine  (see 
page  123)  formed  from  codeine  by  the  mineral  acid. 

Ci7Hi8(CH3)N03  +  HCl  =  CnHnNOa  +  CH3.CI  +  H2O 

Codeine  Apomorphine 

8.  Mecke's  Test. — The  reagent,  consisting  of  selenious  acid 
and  concentrated  sulphuric  acid,^  dissolves  codeine  with  a  blue 
color  quickly  changing  to  emerald  green  and  finally  becoming 
a  permanent  olive  green. 

NARCOTINE 

Narcotine,  C22H23NO7,  crystallizes  in  shining  prisms  or  in  tufts  of  needles 
which  are  nearly  insoluble  in  cold  water  but  readily  soluble  in  boiling  alcohol 
or  chloroform.     Separation  of  alkaloid  from  the  cold  alcoholic  solution  is  almost 

^  See  preparation  of  reagents,  page  315. 


NON-VOLATILE    POISONS  109 

complete.     At  15°  narcotine  dissolves  in  170  parts  of  ether;  31  parts  of  acetic 

ether;  and  22  parts  of  benzene.     Solutions  of  narcotine  are  not  alkaline  nor 

bitter.      In    these    respects    narcotine    is    very 

OCH3  different  from  the  other  opium  alkaloids.     Salts 

I  of  narcotine  do  not  crystallize,  their  stability  is 

A\  slight  and  their  solutions  react  acid.     Salts  with 

HC      C.OCH3    volatile  acids  are  decomposed,  when  their  solutions 

I        II  are  evaporated,  with  separation  of  narcotine.     So- 

■^   /I  dium  acetate  precipitates  free  narcotine  from  its 

X  solution  in  hydrochloric  acid. 

I        I  Constitution. — Narcotine  is  a  monacid,  tertiary 

HC —  O  base  and  as  such  combines  with  i  mol.  of  CH3.I, 

rw  n  r      rw  forming  narcotine    methyl    iodide    (C22H23NO7.- 

CH3I).      This  compound  is  formed  at  ordinary 


,0 — C     C      N.CH3       temperatures    but    the    reaction  is   hastened    by 

H2C<('  I        II        I  heat.     Narcotine  heated  with  hydriodic  acid  loses 

^"^  ^  ^^2  3  methyl   groups   which   form    CH3I.      The    al- 

/    ^  akloid    must    therefore    contain     3  'methoxyls, 

H     H2  sCCHsO-)  groups,  in  the  molecule.     Heated  with 

water  to  140°,  with  dilute  sulphuric  acid,  or  even 

with   barium   hydroxide    solution   narcotine  is   hydrolyzed  into   nitrogen-free 

opianic  acid  and  into  the  basic  and  consequently  nitrogenous  hydrocotarnine: 

C22H23NO7  4-  H2O  =  C10H10O5  +  C12H15NO3 
Narcotine.  Opianic         Hydrocotarnine. 

acid. 

By  oxidative  cleavage,  that  is,  by  treatment  of  narcotine  with  such  oxidizing 
agents  as  nitric  acid,  manganese  dioxide  and  sulphuric  acid,  lead  dioxide  and 
ferric  chloride,  cotarnine  and  opianic  acid  are  the  products: 

C22H23NO7  +  (H2O  +  O)  =  C10H10O5  +  C12H16NO4 
Narcotine  Opianic  Cotarnine 

acid  * 

Evidently  these  cleavage  products  show  that  this  alkaloid  is  made  up  of  two 
complexes,  one  nitrogen-free  and  the  other  containing  nitrogen.  The  chemical 
constitution  of  these  cleavage  products  has  been  determined  and  is  expressed  by 
the  following  formulae: 

H  CH3O 

C  I      Ho 

/\  c    c 

CH3O— C      CH 


I        II  /O— C      C      N— CH3 
CH3O— C       C— C=0                              H2C<  1        II        I 

\/      H  \0— C      C      CH2 

c  \/\/ 

I  c     c 

HO— C=0  H      H2 

Opianic  Acid  Hydrocotarnine 


110  DETECTION   OF  POISONS 

CH3O  O 

!        / 

C      C— H 
/O— C     C       NH.CH3 

H2C/     I    II     I 

^O— C      C       CH2 


c    c 

H     H2 

Cotarnine 

On  the  basis  of  these  results,  Roser  and  Freund  have  proposed  the  structural 
formula  for  narcotine  given  above.  They  consider  the  constitution  of  this  alka- 
loid as  definitely  settled.  If  the  formula  of  narcotine  is  compared  with  that  of 
hydrastine  (see  page  112),  a  great  similarity  in  structure  will  be  seen.  In  fact 
narcotine  is  a  methoxylized  hydrastine. 

Detection  of  Narcotine 

Narcotine  is  so  feebly  basic  that  chloroform  will  extract  the 
alkaloid  completely  from  an  aqueous  tartaric  acid  solution. 
Consequently  its  separation  from  the  rest  of  the  opium  alkaloids 
as  well  as  from  other  alkaloids  is  easy.  Naturally  ether  or 
chloroform  will  also  extract  narcotine  from  an  aqueous  alkaline 
solution.  The  alkaloid  as  it  comes  from  its  ether  solution  is 
usually  a  slightly  colored,  varnish-like  residue  which  hardens 
after  a  time  to  a  mass  of  radiating  crystals.  Narcotine  is 
precipitated  from  its  hydrochloric  or  sulphuric  acid  solution 
by  iodo-potassium  iodide,  phospho-molybdic  acid,  potassium 
mercuric  iodide,  potassium  bismuthous  iodide  even  in  consider- 
able dilution  (i  :5ooo). 

1.  Sulphuric  Acid  Test. — Dissolved  with  stirring  in  concen- 
trated sulphuric  acid,  narcotine  produces  a  greenish  yellow  color 
which  gradually  changes  to  reddish  yellow  and  finally  after 
several  days  to  raspberry  red. 

2.  Dilute  Sulphuric  Acid  Test. — A  solution  of  narcotine  in 
dilute  sulphuric  acid  (i  :5),  evaporated  on  the  water-bath  in  a 
porcelain  dish  or  over  a  very  small  flame,  has  a  reddish  yellow 
color,  changing  with  stronger  heat  to  crimson  red.  As  the  acid 
begins  to  evaporate,  blue-violet  streaks  radiate  from  the  margin 
and  finally  the  entire  liquid  has  a  dirty  red  violet  color  (Dragen- 
dorff's  reaction) .    The  same  color  changes  appear,  if  the  yellow- 


NON-VOLATILE   POISONS  111 

ish  solution   of  narcotinc  in   concentrated  sulphuric   acid   is 
heated  very  carefully. 

3.  Froehde's  Test. — This  reagent  dissolves  narcotine  with  a 
greenish  color.  If  concentrated  Froehde's  reagent  is  used,  the 
green  color  changes  immediately  to  cherry  red,  especially  upon 
application  of  gentle  heat.     This  color  is  quite  persistent. 

4.  Couerbe's  Test. — Dissolve  narcotine  in  cold  concentrated 
sulphuric  acid  and  mix  a  trace  of  nitric  acid  with  this  solution 
after  1-2  hours.  A  red  color  will  appear  and  gradually  become 
more  and  more  pronounced. 

Erdmann's  reagent  gives  the  same  color  change. 

5.  Wangerin's  Test.^ — Place  a  mixture  of  o.oi  gram  of 
narcotine  with  20  drops  of  pure  concentrated  sulphuric  acid 
and  1-2  drops  of  i  per  cent,  cane  sugar  solution  upon  a  watch 
glass  and  heat  upon  the  water-bath  with  stirring  about  i  minute. 
At  first  the  solution  has  a  greenish  yellow  color  which  passes 
through  yellow,  brownish  yellow,  brown  and  brown-violet  into 
an  intense  blue-violet. 

The  intensity  of  this  color  increases  somewhat  upon  standing 
and  the  blue-violet  color  persists  several  hours. 

Applied  to  apomorphine,  atropine,  brucLne,  quinine,  codeine,  caffeine,  hydras- 
tine,  morphine,  physostigmine,  pilocarpine  and  strychnine,  this  test  gives 
solutions  that  are  colorless  or  nearly  so.  Only  the  morphine  solution  after  a 
while  has  a  pale  pink  color.  Coniine  and  narcotine  have  a  light  yellow  color; 
narceine  chestnut-brown;  and  picrotoxin  salmon  color  to  pale  pink. 

Colchicin,  digitalin  and  veratrine  behave  toward  this  reagent  as  toward  pure 
concentrated  sulphuric  acid  without  the  addition  of  the  small  quantity  of  sugar. 

In  this  test  1-2  drops  of  i  per  cent,  aqueous  furfurol  solution  may  be  substi- 
tuted for  the  sugar  solution.  From  yellow,  brown,  olive  and  other  colors  there 
finally  emerges  a  deep,  clear,  dark  blue.  The  brilliancy  of  this  color  increases 
somewhat  on  standing.  After  several  hours  there  is  a  gradual  change  to  a  pure 
green  color.  For  the  detection  of  traces  of  narcotine  (o.ooi  gram)  use  a  i  per 
cent,  sugar  solution. 

6.  Selenious  Acid-Sulphtiric  Acid  Test. — This  reagent  dis- 
solves narcotine  with  a  greenish  steel-blue  color  w^hich  after  a 
time  becomes  cherry-red.  Heat  immediately  discharges  the 
cherry-red  color. 

^  Pharmazeutische  Zeitung,  48,  607    (1903). 


112  DETECTION   OP   POISONS 


HYDRASTINE 


Hydrastine,  C21H21NO6,  occurs  together  with  berberine,  C20H17NO4,  and  cana- 

dine,  C20H21NO4,  in  hydrastis  root,  the  root  of  Hydrastis  canadensis,  to  the 

amount  of  1.5  per  cent,  and  more.     The  fluid   ex- 

O.CH3  tract  prepared  from  this  root  and  used  in  medicine 

1  contains  2-2.5  per  cent,  of  hydrastine. 

/?-\      .  Preparation. — Extract    hydrastis    root   with   hot 

HC       C.OCH3   water  containing  acetic  acid.      Filter  the  solution, 

I        II  evaporate  to  a  thin  extract  and  add  ^  vols,  of  dilute 

HC       C  CO 

^  y,  '  sulphuric    acid    (1:5).      Nearly  all  the  berberine 

Q  separates  out  in  fine  yellow  crystals  as  acid  sul- 

I  phate,    C20H17NO4.H2SO4.     Precipitate   hydrastine 

HC O  from   the   mother-liquor  of   berberine  sulphate  by 

I  means  of  ammonium  hydroxide  solution  and  purify 

~^\  /\  the   alkaloid  by  crystallization  from  acetic  ether  or 

.0— C      C      N.CH3     alcohol.     Hydrastine    crystallizes  from   alcohol   in 
H2CX  I        II        I  rhombic  prisms  melting   at   132°.     It  is  nearly  in- 

S(    y^     C-n.2         soluble  in  water  but  freely  soluble  in  hot  alcohol, 
Q      Q  benzene  or  chloroform.      This  alkaloid  has  a  bitter 

H     H2  taste   and  its   solutions   are   alkaline.     Hydrastine 

solutions  are  optically  active.  In  chloroform  this 
alkaloid  is  laevo-rotatory,  whereas  in  dilute  hydrochloric  acid  it  is  dextro- 
rotatory. 

Constitution. — The  constitution  of  hydrastine  is  entirely  analogous  to  that  of 
narcotine  (see  page  109).  On  oxidation  with  dilute  nitric  acid  hydrastine  gives 
opianic  acid  and  hydrastinine : 

C21H21NO6  +  (H2O  +0)  =  CioHiops  +  CnHisNOs 
Hydrastine  Opianic  acid        Hydrastinine 

Hydrastine  is  a  monacid  base  which  is  shown  to  be  a  tertiary  base  by  its 
behavior  toward  alkyl  iodides,  for  example,  with  CH3I  it  forms  hydrastine  methyl 
iodide,  C21H21NO6.CH3I,  which  crystallizes  in  needles.  Hydrastine  contains 
two  methoxyl  groups,  because  when  heated  with  hydriodic  acid  according  to 
Zeisel's  method  two  such  groups  are  removed. 

Since  the  chemical  nature  of  opianic  acid  has  long  been  known,  the  only 
problem  is  the  explanation  of  the  nature  of  hydrastinine,  the  other  cleavage 
product.  The  constitution  of  hydrastinine,  as  well  as  that  of  many  other  alka- 
loids, has  been  determined  by  A.  W.  Hofmann's  method  of  exhaustive  methyla- 
tion.^     Hydrastinine  (I)  is  a  secondary  base  which  forms,  when  heated  with  an 

1  When  the  nitrogen  of  an  organic  base  becomes  quinquevalent,  it  is  more  sub- 
ject to  change.  Hofman  (Liebig's  Annalen,  78,  263  (1851)  showed,  for  example, 
that  tetra-ethyl-ammonium  hydroxide  breaks  up  on  heating  into  triethylamine, 
ethylene  and  water: 


C2H5 
C2H5 
C2H5 


^  TT      \  CH2  C2H5\ 

•^^  r'>N-OH  =    II       +  CjHs^N  +  H2O. 


p    TT    /  CH2  C2H6- 

Nitrogen  in  alkaloids  on  treatment  with  an  alkyl  haloid  (e.g.,  CH3I)  combines 
with  it  in  many  instances,  forming  compounds  having  a  structure  analogous 


NON-VOLATILE   POISONS  113 

excess  of  CH3I,  hyclrastinine  hydriodide  and  trimethyl-hydrastyl-ammonium 
iodide  (II).  Heated  with  alkalies,  this  ammonium  iodide  is  decomposed  into 
trimethylamine,  hydriodic  acid  and  nitrogen-free  hydrastal  (III).  The  latter 
on  oxidation  gives  hydrastic  acid  (IV)  which  was  recognized  as  the  methylene 
ether  of  nor-meta-hcmipinic  acid  (V) : 

/CH:0  +  2CH3I  = 

(I)  (CH202)CoH2< 

\CH2.CH2.NH.CH3 

/CH :  O 

(CH202)C6H2<  V 

\CH2.CH2.N(CH3)3l 

Hydrastinine  Trimethyl-hydrastyl- 

ammonium    iodide 

/CH:0 

(II)  (CH202)C6H2< 

\CH2.CH2.N(CH3)3l  +  KOH  = 
/CH:0 
(CH202)C6H2<  +  Kl-f  H2O  +  N(CH3), 

^CH:CH2 

Hydrastal 

/CH:0        •  /COOH 

(III)  (CH202)C6H2<  Oxidized  =  (CH202)C6H2<  (IV) 

\CH:CH2  ^COOH 

Hydrastic  acid 


Hydrastic  acid  and  nor-meta-hemipinic  acid  are  identical.     The  latter  has  the 

structure  (V): 

H 

C 

/\ 

(V)  H2C< 

-C      C.COOH 

1        1 

-C      C.COOH 

\/ 

c 

H 

Nor-meta-hemipinic  acid 

From  these  and  other  relations  it  has  been  determined  that  cotarnine  is  a 

methoxy-hydrastinine : 

H 

CH3.O      H 

H        / 

1        1 

c     c=o 

c    c=o 

/\/ 

/\/ 

/O— C      C       NH.CH3 

H2C<         1        II        1 
\0— C      C      CH2 

/O— C      C       NH.CH3 

H2C<          1        11        1 
^0— C      C       CHo 

\/\/ 

\/\/ 

c    c 

c    c 

H     H2 

H     Ho 

Hydrastinine 

Cotarnine 

The  alkaloid  narcotine  is  a  methox^'-hj^drastine  (see  page  109). 

to  that  of  tetra-ethyl-ammonium  hydroxide.  This  process  is  called  "exhaustive 
meth3dation."  Upon  decomposition  these  deri\atives  yield  products  which 
often  throw  light  upon  the  structure  of  the  alkaloid. 


114  DETECTION   OF  POISONS 

Detection  of  Hydrastine 

1.  Concentrated  Sulphuric  Acid  dissolves  hydrastine  without 
color  but  upon  being  gently  warmed  the  solution  becomes  violet. 

2.  Froehde's  Reagent  dissolves  hydrastine  with  a  green  color 
which  gradually  changes  to  brown. 

3.  Mandelin's  Reagent  dissolves  hydrastine  with  a  rose- 
red  color  which  immediately  changes  to  orange-red  and  gradu- 
ally fades. 

4.  Fluorescence  Test. — Dissolve  hydrastine  in  dilute  sul- 
phuric acid,  shake  vigorously  and  add  drop  by  drop  very 
dilute  potassium  permanganate  solution.  Hydrastinine  is 
formed  and  the  solution  shows  a  beautiful  blue  fluorescence. 

.  The  ether  extract  of  the  alkaline  solution  on  evaporation 
leaves  hydrastine  in  a  crystalline  condition. 

QUININE 

Quinine,  C20H24N2O2,  is  precipitated  amorphous  and  anhydrous  from  solutions 
of  its  salts  by  caustic  alkalies,  alkaline  carbonates  or  ammonia.     On  standing, 
jj  however,  it  gradually  becomes  crystalline,  forming  a 

C  hydrate   with   3    molecules  of   water  of  hydration. 

/|\  There  are  also  other  hydrates  of  quinine.      Anhy- 

^2p     I     CH.CH:CH2    drous  quinine  melts  at  173°;  the  trihydrate  at  57°. 
I     ^  An  ether  solution  on  evaporation  usually  deposits 

HO.C     CH2CH2  this  alkaloid  as  a  resinous,  or  varnish-like,  amorphous 

^  I  /  residue.       Quinine  is  soluble  in  about  2000  parts  of 

I  cold  and  700  parts  of  boiling  water;  and  freely  solu- 

JqH  ble  in  alcohol,  ether  or  chloroform.     Solutions  of 

I       H  quinine  in  sulphuric,  acetic  or  tartaric  acid  exhibit  a 

C      C  beautiful  blue  fluorescence.     In  the  case  of  the  sul- 

/f  \<  ^  /-^o■rT        phate  this  fluorescence  is  distinctly  visible  in  a  dilu- 
I        II       I  tion  or  I  :  100,000. 

HC      C      CH  Hydrochloric,  hydrobromic  and  hydriodic  acid  do 

\X\^  not  give  fluorescent  solutions  of  quinine.    These  acids 

■^^      ji  even  discharge  the  fluorescence,  if  added  to  a  fluor- 

escent quinine  solution. 
Constitution. — Quinine  is  a  diacid,  ditertiary  base,  the  salts  of  which  with 
I  and  2  equivalents  of  acid  are  usually  crystalline.  The  salts  with  i  equivalent 
of  acid  are  the  more  stable.  Quinine  hydrochloride,  C20H24N2O2.HCI.2H2O, 
used  in  medicine,  crystallizes  in  long  delicate  tufts  of  needles.  The  diter- 
tiary character  of  quinine  is  shown  by  the  fact  that  it  unites  with  2  mole- 
cules of  methyl  iodide,  for  example,  to  form  quinine  dimethyliodide, 
C20H24N2O2.2CH3I.  Quinine  must  contain  an  hydroxyl  group,  since  it  can 
form  a  mono-benzoyl    and  a  mon-acetyl-quinine.     Moreover    one    methoxyl 


NON-VOLATILE   POISONS 


115 


group  has  been  found  in  the  quinine  molecule.  The  difference  empirically 
between  cinchonine,  C10II22N2O,  and  quinine,  C20H24N2O2,  is  CII2O.  Every 
investigation  of  these  substances  has  shown  that  quinine  is  a  methoxy-cincho- 
nine.  For  example,  on  oxidation  with  chromic  acid,  cinchonine  gives  cinchonic 
acid  which  was  recognized  as  quinolinc  7-carboxyIic  acid;  whereas  quinine  under 
the  same  conditions  gives  quinic  acid,  or  p-methoxy-cinchonic  acid: 


coohCt) 

H.       1 

c     c 

HC     C      CH 

(P) 

C00II(7) 
H      1 

c    c 

CHaO.C      C      CH 

HC      C      CH 

C     N 
H 

Cinchonic  acid 

HC      C      CH 

C     N 
H 

Quinic  acid 

Both  alkaloids  on  oxidation  also  give  the  nitrogenous  compounds  mero- 
quinene,  cincholoiponic  acid  and  loiponic  acid.  Consequently  there  is  no 
doubt  that  cinchonine  and  quinine  contain  two  nitrogenous  nuclei,  one  of  which 
is  a  quinoline  complex.  The  second  nucleus  is  connected  with  the  latter  in  the 
7-position,  as  the  formation  of  cinchonic  and  quinic  acids  shows.  Meroquinene, 
cincholoiponic  acid  and  loiponic  acid,  derived  by  oxidation  with  chromic  add 
from  the  so-called  "second  half"  of  the  cinchonine  and  quinine  molecules,  form 
a  continuous  series  of  oxidation  products,  since  meroquinene  can  be  oxidized  to 
cincholoiponic  acid  and  the  latter  to  loiponic  acid.  The  following  formulae  best 
explain  the  chemical  behavior  of  these  three  compounds : 


CH2.COOH 
CH 


CH2.COOH 

I 
CH 


COOH 

1 
CH 


H2C     CH— CH  :  CH2 

H2C     CH2 

\/ 
N 
H 

Meroquinene 


H2C     CH.COOH        H2C     CH.COOH 


H2C      CH2 

\/ 

N 
H 

Cincholoiponic  acid 


H2C       CH2 


N 
H 
Loiponic  acid 


The  structural  formula  already  given  for  quinine  was  proposed  by  W.  Koenigs^ 
and  is  based  on  the  results  of  his  own  experiments  as  well  as  on  those  of  \V.  V. 
Miller  and  of  Skraup.  Cinchonine  has  hydrogen  in  place  of  the  methoxyl  group 
in  the  quinoline  nucleus;  otherwise  the  two  alkaloids  are  identical  in  structure. 


Detection  of  Quinine 

Ether,  benzene  or  chloroform  will  extract  quinine  from  an 
aqueous  alkaline  solution.     Ether  on  evaporation  deposits  the 

'  Meroquinene  and  the  Structure  of  the  Cinchona  Alkaloids;  Annalen  der 
Chemie  und  Pharmazie  347,  147  (1906). 


116  DETECTION   OF   POISONS 

alkaloid  as  a  resinous,  amorphous  varnish  in  which  its  presence 
may  be  recognized  by  the  following  tests: 

1.  Fluorescence  Test. — Dissolve  the  residue  from  the  ether 
extraction  of  the  alkaline  solution  in  a  little  dilute  sulphuric 
acid.  If  quinine  is  present,  this  solution  will  exhibit  blue 
fluorescence. 

2.  Thalleioquin  Test. — Dissolve  quinine  in  a  few  drops  of 
very  dilute  acetic  acid  and  add  5-10  drops  of  saturated  chlorine 
water.  The  colorless  solution  has  a  faint,  blue  fluorescence. 
Excess  of  ammonium  hydroxide  solution  will  produce  an  emer- 
ald green  color.  A  solution  containing  considerable  quinine 
will  give  a  green  precipitate.  This  precipitate  (thalleioquin) 
is  always  an  amorphous  substance,  the  composition  of  which 
has  not  been  determined.  It  is  soluble  in  alcohol  and  chloro- 
form but  not  in  ether. 

E.  Polacci  recommends  the  following  procedure  for  the  thal- 
leioquin test.  Gradually  heat  quinine  (about  o.oi  gram)  to 
boiling  with  a  little  lead  dioxide  (Pb02) ,  2-3  cc.  of  water  and  2 
drops  of  dilute  sulphuric  acid.  Let  the  solution  settle  and 
either  decant  or  filter.  Finally,  carefully  add  5-6  drops  of 
ammonium  hydroxide  solution  as  a  top  layer.  A  fine  green 
ring  will  appear  at  the  zone  of  contact. 

Interferences  with  the  Thalleioquin  Test. — Antipyrine  interferes  with  this  test. 
Mixtures  of  i  per  cent,  solutions  of  antipyrine  and  quinine  give  finally  a  beautiful 
red  instead  of  a  green  color.  This  interference  does  not  cease  until  these  two 
substances  are  in  the  proportion  of  0.25  parts  of  antipyrine  to  5  parts  of  quinine. 
Caffeine  also  interferes  with  the  thalleioquin  test,  when  the  proportion  is  2  parts 
of  quinine  to  3  parts  of  caffeine.  Other  compounds  like  urea  prevent  the  appear- 
ance of  this  color,  whereas  morphine,  pilocarpine,  cocaine,  atropine,  codeine, 
strychnine,  carboUc  acid  and  chloral  hydrate  have  no  effect  upon  the  thalleioquin 
test. 

H.  Fiihneri  has  shown  that  the  thalleioquin  reaction  is  connected  with  the 
p-oxyquinoline  complex.  Chlorine  passed  into  a  solution  of  pure  p-oxy-quino- 
hne  cooled  with  ice  produces  a  white  crystalline  precipitate.  This  substance 
crystallizes  from  petroleum  ether  in  colorless  prisms  or  tabular  crystals  melting  at 
58°.  Structurally  it  is  5,5-dichloro-6-keto-quinoline.  Solutions  of  this  dichloro- 
keto-quinoHne  and  of  its  hydrochloride  are  colored  a  pure  green  or  blue  by  am- 
monium hydroxide.  Fuhner  thinks  5,6-quinohne  quinone  is  probably  formed 
and  gives  the  green  color  with  ammonia. 

1  Berichte  der  Deutschen  chemischen  Gesellschaft  38,  2713  (1905). 


NON-VOLATILE   POISONS  117 


H     H 

H     CU 

H     0 

C      C 

C      C 

C      C 

/\/\ 

/\/\ 

^\/- 

HC      C      C.OH(p) 

HC      C      CO 

HC      C      CO 

1       II        1             "*■ 

1       II        1 

— >      [       II        1 

HC      C      CH 

HC      C      CH 

HC      C      CH 

\/\/- 

\/\/ 

\/\/ 

N      C 

N     C 

N      C 

H 

H 

H 

p-Oxy-quinoline 

5,S-Dichloro-keto- 

S,6-0uinoline 

quinoline 

cjuinone 

3-  Herapathite  Test. — Mix  30  drops  of  acetic  acid,  20  drops 
of  absolute  alcohol  and  i  drop  of  dilute  sulphuric  acid.  Add 
20  drops  of  this  mixture  to  o.oi  gram  of  quinine  and  heat  to 
boiling.  Finally  add  i  drop  of  an  alcoholic  solution  of  iodine 
(i  :io)  or  2  drops  of  o.i  n-iodine  solution.  When  the  solution 
has  stood  for  some  time,  green  leaflets  with  a  metallic  luster  will 
form.  This  is  an  iodine  compound  of  quinine  called  "Hera- 
pathite," having  the  constant  composition 

4C20H24N2O2.3H2SO4.2HI.3H2O. 

This  substance  can  be  recrystallized  from  boiling  alcohol. 
Herapathite  crystals  are  pale  olive-green  by  transmitted 
light  but  by  reflected  light  they  have  a  beautiful,  cantharidin- 
green,  metallic  luster. 

Caustic  alkalies,  ammonia,  sulphurous  acid  and  hydrogen  sulphide  decompose 
herapathite.  A.  Christensen  recommends  keeping  on  hand  the  following  reagent 
for  the  herapathite  test: 

Parts 
Iodine  i 

Hydriodic  acid  (50%)  i 

Sulphuric  acid  o  .8 

Alcohol  (70%)  50 

Add  a  few  drops  of  this  reagent  to  the  alcoholic  solution  to  be  tested  for  quinine. 

4.  Hirschsohn's  Test.^ — If  i  drop  each  of  2  per  cent,  hydro- 
gen dioxide  and  10  per  cent,  copper  sulphate  solution  are  added 
to  a  neutral  solution  of  quinine  hydrochloride  or  sulphate  at 
boiling  temperature,  a  more  or  less  intense  raspberry  red  color 
will  appear.  This  color  soon  passes  through  blue-\dolet  into 
blue  and  after  a  time  into  green.  A  quiiune  solution  (i  :  10,000) 
will  still  give  a  distinct  red-violet  color. 

^  Pharmazeutische  Zentral-HaUe  43,  367  (1902). 


118  DETECTION   OF  POISONS 

Excess  of  acid  as  well  as  of  alcohol  interferes  with  this  test.  The  behavior 
of  a  solution  of  aloes  toward  this  test  is  similar  to  that  of  quinine. 

Of  the  alkaloidal  reagents  potassium  bismuthous  iodide  is 
especially  recommended  as  a  precipitant  of  quinine.  With 
quinine  sulphate  solutions  this  reagent  produces  precipitates 
having  an  intense  yellowish  red  color.  Shaken  with  sodium 
hydroxide  solution  this  precipitate  is  decomposed  and  unaltered 
quinine  can  be  obtained  by  extraction  with  ether  and  evaporation 
of  the  ether  solution.  H.  Thorns^  has  made  use  of  this  reac- 
tion in  the  quantitative  separation  of  quinine  from  mixtures. 

CAFFEINE 

Since  caffeine  (see  page  79)  is  a  weak  base,  ether  will  extract 
only  a  little  of  the  alkaloid  from  the  tartaric  acid  solution.  The 
greater  part  will  be  in  the  ether  extract  of  the  alkaline  solution. 
Ether  usually  deposits  caffeine  in  white,  shining  needles  ar- 
ranged in  clusters.  Caffeine  dissolves  in  ether  with  some 
difficulty  and  the  alkaline  solution  should  be  extracted  several 
times.     For  the  tests  characteristic  of  this  alkaloid  see  page  80. 

ANTIPYRINE 

Most  of  the  antipyrine  (see  page  78)  is  obtained  by  extract- 
ing the  alkaline  solution  with  ether.  It  is  usually  purer  from 
the  acid  than  from  the  alkaline  solution  and  frequently  appears 
in  crystalline  leaflets.  Antipyrine  differs  from  most  alkaloids 
in  having  only  a  faintly  bitter  taste  and  in  being  freely  soluble 
in  water.  To  identify  antipyrine,  dissolve  the  ether  residue  in  a 
little  water  and  divide  the  solution  into  two  equal  parts.  Test 
one  portion  with  ferric  chloride  solution  and  the  other  with 
fuming  nitric  acid. 

Detection  of  Antipyrine  in  Urine. — The  color  of  urine  after  administration  of 
antipyrine  is  intensely  yellow  to  blood  red.  Part  of  the  antipyrine  in  the  organ- 
ism appears  in  the  urine  as  oxy-antipyrine-glycuronic  acid  and  another  part  is 
unchanged  and  can  usually  be  detected  directly  in  urine  by  ferric  chloride  solu- 
tion. A  safer  procedure  is  to  add  excess  of  ammonia  to  a  considerable  quantity  of 
urine  and  extract  with  chloroform.     Evaporate  the  solvent,  dissolve  the  residue 

^Berichte  der  Deutschen  pharmazeutischen  Gesellschaft  16,  130  (1906). 


NON-VOLATILE   POISONS  119 

in  a  little  water  and  test  the  filtered  solution  for  antipyrine  with  ferric  chloride 
solution  and  with  fuming  nitric  acid. 

Antipyrine  is  easily  absorbed.  The  urine  may  show  a  reddish  color  even  an 
hour  after  the  drug  has  been  taken  and  give  a  test  with  ferric  chloride  solution. 
The  red  color  disappears  in  about  24  hours  but  the  elimination  of  antipyrine  is 
not  complete  in  that  time.  Its  detection  is  still  possible  after  36  hours.  A  con- 
venient procedure  is  to  add  to  the  urine  as  an  upper  layer  very  dilute  ferric  chlo- 
ride solution.  A  red  ring  will  appear  if  the  urine  contains  antipyrine.  Jonescu^ 
states  that  antipyrine  in  the  human  organism  passes  unchanged  into  the  urine. 
Only  a  small  portion — and  large  doses  of  the  drug  must  have  been  taken — is 
eliminated  in  conjugation  with  sulphuric  acid.  Conjugation  with  glycuronic 
acid^  (see  above)  according  to  Jonescu  does  not  occur  in  the  human  organism. 

PYRAMmONE 

Pyramidone,  or   4-dimethyl-amino-antipyrine,  CisHnNsO,  has  been   exten- 
sively used  in  medicine  of  late  as  an  antipyretic  and  anodyne.     It  is  a  white, 
P  TT  crystalline  powder,  nearly  tasteless  and  readily  soluble 

I  in  water.     It  melts  at  108°.     Its  aqueous  solution  has 

N  a  neutral  reaction.     Ether  removes  only  traces  of 

/^\  pyramidone  from  acid  solution,  but  extracts  it  easily 

v^jis     1^2     6V.W  and  completely  from  alkaline  solution.     Ether  usually 

QH^ Q3^r^4Q N(CH  )      deposits  this  substance  in  fine  needles.     Pyramidone 

is  also  freely  soluble  in  alcohol,  ether,  chloroform  or 
benzene.  It  is  a  strong  reducing  agent  and  in  this  respect  difJers  from  antipyrine. 
For  example,  pyramidone  will  reduce  gold  chloride  even  in  the  cold,  whereas 
antipyrine  and  tolypyrine  require  heat. 

Preparation. — Antipyrine  dissolved  in  concentrated  acetic  acid  is  converted  by 
treatment  with  potassium  nitrite  into  nitroso-antipyrine  which  appears  as  green 
crystals.  This  compound  dissolved  in  alcohol  may  be  reduced  by  zinc  and  acetic 
acid  to  amino-antipyrine.  The  latter,  dissolved  in  methyl  alcohol  and  treated 
with  methyl  iodide  and  potassium  hydroxide,  is  converted  into  dimethylamino- 
antipyrine,  or  pyramidone. 

CeHe  CeHs  CeHs 

I  I  I 

N  N  N 

/\  /\  /\ 

CH3.N     CO  CH3.N     CO  CH3.N      CO-I-2CH3I 

I        I  .    -^  I        I  +4H->  I         I  -^ 

CH3.C=C:H  HOi.NO     CH3.C=C.N0  CH3.C  =C.NH2  2KOH 

Antipyrine  Nitroso-antipyrine  Amino-antipyrine 

^  Berichte  der  Deutschen  pharmazeutischen  Gesellschaft  16,  133  (1906). 

2  Glycuronic  acid,  CeHioO?  =        >C(CH.0H)4C00H,  may  be  regarded  as  a 

O^ 
derivative  of  glucose.  Possibly  it  occurs  in  normal  urine  in  small  quantity  as 
a  conjugated  acid.  After  administration  of  various  alcohols,  aldehydes,  ketones, 
phenols  (chloral  hydrate,  camphor,  phenol,  thymol,  menthol,  borneol),  there 
takes  place  in  the  animal  organism — often  after  oxidation  or  reduction — a  con- 
jugation of  these  substances  with  glycuronic  acid. 


120  DETECTION   OF   POISONS 

Cells 

I 

N 

/\ 
CHg.N     CO 

CH3.C=C.N(CH3)2 

Pyramidone 

Behavior  in  the  Organism. — Human  urine,  if  neutral  or  faintly  acid,  usually 
has  a  bright  purplish  red  color  after  administration  of  pyramidone.  After  stand- 
ing for  some  time  it  will  deposit  a  sediment  consisting  of  red  needles  soluble  in 
ether  or  chloroform  but  especially  in  acetic  ether.  Jaffe^  recognized  this  com- 
pound as  rubazonic  acid,  a  pryazolone  derivative.  Isolation  of  rubazonic  acid 
from  urine  may  be  brought  about  as  foUows.  Acidify  fresh  urine  with  hydro- 
chloric acid  and  let  it  stand  in  an  open  dish.  The  acid  will  appear  as  small  red 
particles.  Ferric  chloride  solution  produces  a  blue -violet  color  in  the  acid  liquid 
filtered  from  rubazonic  acid.  This  filtrate  contains  most  of  the  product  formed 
from  pyramidone  in  animal  metabolism,  namely,  crystalline  antipyryl-urea 
melting  at  about  245°. 

CeHs 

1 
N 

/\ 
CH3.N     CO 

CH3.C=C.NH.CO.NH2 

Antipyryl-urea 


Detection  of  Pyramidone 

1.  Ferric  Chloride  Test. — Ferric  chloride  solution  added  to 
pyramidone  produces  a  blue-violet  color  which  soon  changes  to 
reddish  violet  and  then  disappears. 

2.  Filming  Nitric  Acid  Test. — A  few  drops  of  faming  nitric 
acid,  added  to  a  solution  containing  pyramidone,  give  a  blue  to 
blue-violet  color. 

3.  Bromine  Water  Test. — This  reagent  imparts  a  grayish 
color  to  pyramidone  solutions.  With  concentrated  solutions 
it  produces  an  inky  color. 

4.  Iodine  Test. — Tincture  of  iodine  colors  an  aqueous  pyra- 
midone solution  blue. 

iBerichte  der  Deutschen  chemischen  Gesellschaft  34,  2737  (1901);  and  35, 
289X  (1902). 


NON-VOLATILE   POISONS  121 

C.  Extraction  of  the  Araraoniacal   Solution  with  Ether  and 
Chloroform 

(a)  Ether  Extract. — Apomorphine  and  traces  of  morphine.^ 

(jS)  Chloroform  Extract. — Morphine  and  narccinc.  (It  may 
also  contain  antipyrinc  and  caffeine.^) 

The  aqueous  alkaline  solution  (see  page  8i),  separated 
from  ether,  must  be  tested  further  for  the  substances  under 
a  and  /3. 

Apomorphine  may  be  recognized  by  the  green  color  of  the 
aqueous  acid  solution.  Excess  of  sodium  hydroxide  solution 
causes  oxidation,  especially  if  the  solution  is  exposed  for  any 
length  of  time  to  air,  and  gradually  changes  the  color  to  deep 
purple-red.  Moreover,  the  ether  extracts,  both  of  the  acid 
and  alkaline  solutions,  are  red  or  violet-red  when  apomorphine 
is  present.  Solutions,  examined  by  the  Stas-Otto  method,  not 
having  these  characteristics,  need  not  be  tested  for  apomorphine. 
In  that  case  proceed  at  once  with  the  morphine  and  narceine 
tests. 

To  extract  apomorphine,  morphine  and  narceine  with  the 
proper  solvent,  the  aqueous  solution  separated  from  ether,  which 
is  alkaline  from  sodium  hydroxide  solution  (see  page  8i),  must 
be  rendered  alkaline  with  ammonium  hydroxide  solution. 
First  acidify  the  solution  with  dilute  hydrochloric  acid  (test 
with  blue  litmus  paper)  and  then  add  ammonium  hydroxide 
solution  until  alkaline. 

(a)  If  there  is  any  indication  of  apomorphine,  first  extract  the 
ammoniacal  solution  repeatedly  with  ether  and  then  several 
times  with  hot  chloroform  for  the  morphine  and  narceine  tests. 

(i8)  If  there  is  no  indication  of  apomorphine,  extract  the 
ammoniacal  solution  several  times  direct  with  hot  chloroform 
(see  below). 

^  Ether  dissolves  traces  of  freshl}'  precipitated,  amorphous  morphine. 

^  Antipyrine  and  caflfeine,  though  freely  soluble  in  chloroform,  dissolve  with 
diflSculty  in  ether.  The  latter  solvent  frequentl}'  fails  to  extract  these  substances 
completely  from  aqueous  solution.  They  wiU  then  appear  in  the  chloroform 
extract. 


(i)    (9) 

H     H2    CH3 

C      C      N 

/\/\/\ 
HC      C      CH  CHi 

1        II       1        1 
(3)H0.C      C      C      CH; 

C      C      C(8) 

(4)H0     HC     CH 

\/ 
C 
H 

122  DETECTION   OF   POISONS 

APOMORPHINE 

Constitution. — ^Apomorphine,  C17H17NO2,  is  a  monacid,  tertiary  base  with  two 
phenol  hydroxyl  groups.  According  to  R.  Pschorr^  it  has  the  structural  formula 
here  given. 

Properties.— Apomorphine  is  an  amorphous  base 
readily  soluble  in  alcohol,  ether,  benzene  or  chloro- 
form and  colored  green  in  contact  with  air.  Aqueous 
and  alcoholic  apomorphine  solutions,  originally  color- 
CH  CH2  less,  soon  turn  green  in  the  air  from  oxidation.  Solu- 
tions of  apomorphine  thus  changed  by  oxidation  are 
emerald  green.  Ether  and  benzene  solutions  are 
purpHsh  violet;  those  in  chloroform  blue- violet.  Be- 
ing phenoUc  in  character,  apomorphine  resembles 
morphine  in  its  solubility  in  sodium  hydroxide  solu- 
tion. Alkaline  solutions  of  the  alkaloid  absorb  oxy- 
gen from  the  air  and  become  brown  or  even  black 
in  color.  Apomorphine  differs  from  morphine  in 
being  more  soluble  in  water  and  in  alcohol,  but  especially  in  being  soluble  in 
ether,  benzene  and  cold  chloroform  in  which  morphine  is  almost  insoluble. 

Formation  and  Preparation. — Sulphuric,  hydrochloric,  phos- 
phoric and  oxalic  acids,  the  alkalies  and  zinc  chloride  have 
mainly  a  dehydrating  action  upon  morphine  and  convert  it 
into  apomorphine: 

C17H19NO3  =  H2O  +  C17H17NO2 

M  orphine  Ap  o  morphine 

Codeine,  the  methyl  ether  of  morphine,  also  gives  apomor- 
phine when  heated  at  140°  with  concentrated  hydrochloric  acid. 

CnHiaNOaCOCHa)  -|-  HCl  =  H2O  -1-  CH3CI  -|-  C17H17NP2 

Codeine  Apomorphine 

Apomorphine  is  prepared  by  heating  morphine  (i  part)  with 
concentrated  hydrochloric  acid  (20  parts)  for  3  hours  in  an 
autoclave  at  130-150°. 

(a)  Detection  of  Apomorphine  in  the  Ether  Extract 

Ether  will  not  extract  apomorphine  from  a  solution  contain- 
ing tartaric  acid  but  will  dissolve  its  colored  oxidation  products. 
This  solvent  behaves  similarly  toward  solutions  of  this  alkaloid 
in  sodium  or  potassium  hydroxide  solutions.     Ether  or  chloro- 

1  Berichte  der  Deutschen  chemischen  Gesellschaft  39,  3124  (1906);  and  40, 
1984  (1907). 


NON-VOLATILE   POISONS  123 

form  will  extract  apomorphine  only  from  a  solution  alkaline 
with  ammonium  hydroxide.  Ether  solutions  of  apomorphine 
usually  deposit  a  greenish  residue.  A  characteristic  of  this 
alkaloid  is  its  strong  reducing  action.  For  example,  it  will  re- 
duce iodic  acid  with  liberation  of  iodine  and  produce  a  purple 
color  with  gold  chloride.  Apomorphine  gives  the  following 
tests : 

1.  Sulphuric  and  Nitric  Acids. — Concentrated  sulphuric  acid 
dissolves  apomorphine  without  color.  Addition  of  a  drop  of 
concentrated  nitric  acid  to  such  a  solution  produces  an  evan- 
escent violet  color  that  soon  changes  to  blood  red  and  finally  to 
yellowish  red.  With  concentrated  nitric  acid  alone  this  alka- 
loid gives  a  violet-red  color  that  soon  becomes  red-brown  and 
finally  brownish  red. 

2.  Pellagri's  Test. — Dissolve  apomorphine  in  dilute  hydro- 
chloric or  sulphuric  acid  and  first  add  acid  sodium  carbonate  in 
excess.  Then  add  drop  by  drop  1-3  drops  of  an  alcoholic  iodine 
solution  and  shake  for  several  minutes.  The  solution  will  have 
a  blue-green  or  emerald  green  color.  Extract  with  a  Httle  ether 
and  the  solvent  will  become  violet,  whereas  the  aqueous  solution 
will  remain  green. 

3.  Froehde's  Test. — This  reagent  dissolves  pure  apomorphine 
with  a  green  color.  If  the  alkaloid  has  been  acted  upon  by  air 
to  any  extent,  the  color  is  violet. 

4.  Wangerin's^  Test. — Prepare  a  fresh  solution  of  apomor- 
phine hydrochloride  (about  i  per  cent.).  Add  4  drops  of  potas- 
sium dichromate  solution  (0.3  per  cent.)  to  i  cc.  of  this  solution 
and  shake  for  about  i  minute.  The  solution  will  have  an  in- 
tense dark  green  color.  Then  add  10  cc.  of  acetic  ether  and 
shake  again.  This  solvent  will  become  violet.  Finally  add 
from  a  pipette  about  5  drops  of  stannous  chloride  solution^ 
(i  per  cent.)  and  shake  well.  The  color  of  the  acetic  ether  layer 
will  change  to  green  and,  upon  further  addition  of  a  few  drops  of 

^  Pharmazeutische  Zeitung  47,  599  and  739-740  (1902). 

2  Prepare  this  reagent  as  follows : 

Crystallized  stannous  chloride  (SnCl2.2H20)         i  gram 
Hydrochloric  acid  (25  per  cent.)  50  cc. 

Water  50  cc. 


124  DETECTION   OF   POISONS 

potassium  dichromate  solution,  the  acetic  ether  will  again  be- 
come violet"  If  lo  cc.  of  chloroform  are  substituted  for  acetic 
ether  in  this  test,  the  oxidation  product  of  apomorphine  will  im- 
part the  same  violet  color  to  the  chloroform.  But  if  stannous 
chloride  solution  is  added  carefully,  the  color  will  change  to  pure 
indigo  blue  and  persist  upon  further  agitation  with  potassium 
dichromate  solution. 

5.  E.  Schmidt's  Tests.  ^ — (a)  A  drop  of  very  dilute  ferric 
chloride  solution  (i  :  100)  will  color  10  cc.  of  an  aqueous  apo- 
morphine hydrochloride  solution  blue  even  in  a  dilution  of 
I  :  10,000. 

(b)  Shake  10  cc.  of  the  same  apomorphine  hydrochloride 
solution  with  i  cc.  of  chloroform.  Then  render  alkaline  with 
sodium  hydroxide  solution  and  at  once  shake  with  air.  The 
aqueous  solution  becomes  evanescent  violet  in  color  and  the 
chloroform  blue. 

((3)  Examination  of  the  Chloroform  Extract 

Preliminary  Morphine  Test.' — As  a  preliminary  test  for  mor- 
phine, acidify  a  small  portion  of  the  aqueous  alkaline  solution 
separated  from  ether  (see  page  81)  with  dilute  sulphuric  acid, 
add  iodic  acid  solution  and  extract  with  a  little  chloroform.  If 
the  latter  has  a  violet  color  from  dissolved  iodine,  morphine 
may  be  present.  But  a  final  conclusion  regarding  the  presence 
of  morphine  must  not  be  drawn  from  a  positive  test,  since  there 
are  many  other  organic  substances  besides  this  alkaloid  that 
will  reduce  iodic  acid.^  This  is  a  delicate  preliminary  test  for 
morphine  and  that  is  its  only  value.  If  it  is  negative,  morphine 
is  probably  absent. 

To  detect  morphine  and  narceine  positively,  render  the  aque- 
ous solution  alkaline  with  ammonium  hydroxide  and  extract  at 
once  as  already  directed  (see  page  121)  with  considerable  hot 
chloroform^  in  a  capacious  flask.     Separate  the  two  liquids  as 

^  Apotheker-Zeitung  23,  657  (1908). 

^  In  testing  animal  matter  that  contained  no  morphine,  the  author  has  repeat- 
edly obtained  extracts  that  strongly  reduced  iodic  acid. 

^  C.  Kippenberger  (Zeitschrift  fiir  analytische  Chemie  39,  201,  290)  uses 
chloroform,  containing  10  per  cent,  of  alcohol  by  volume,  to  extract  morphine. 


NON-VOLATILE    POISONS  125 

usual  in  a  separatory  funnel.  Several  extractions  of  the  aque- 
ous solution  with  fresh  portions  of  hot  chloroform  are  necessary 
because  of  the  slight  solubihty  of  morphine  even  in  boiling 
chloroform.  Should  the  chloroform  and  the  aqueous  solution 
form  a  refractory  emulsion  that  will  not  separate,  add  a  few 
drops  of  alcohol,  set  the  flask  on  a  warm  but  not  boiUng  water- 
bath  and  carefully  turn  the  flask. from  time  to  time.  This 
procedure  usually  causes  the  immediate  separation  of  the  two 
liquids.  Place  the  combined  chloroform  extracts  in  a  dry  flask, 
add  a  few  crystals  of  dry  sodium  chloride  or  anhydrous  sodium 
sulphate  to  remove  adherent  water,  pour  the  chloroform  when 
clear  through  a  dry  filter  and  evaporate  in  not  too  large  a  glass 
dish  placed  upon  a  warm  water-bath.  The  chloroform  may  also 
be  filtered  directly  into  the  dish  as  fast  as  it  evaporates.  If  the 
residue  is  bitter  and  can  be  scraped  together  with  a  platinum 
spatula  or  a  pocket  knife,  test  for  morphine  and  narceine.^  In 
testing  for  morphine  use  Froehde's,  Husemann's  and  Pellagri's 
tests  as  well  as  those  given  by  formalin-sulphuric  acid  and  iodic 
acid.  The  presence  of  morphine  is  not  established  unless  all 
these  morphine  tests  give  positive  results.  If  the  quantity  of 
the  residue  from  chloroform  permits,  test  for  morphine  with 
ferric  chloride  solution.  This  test  is  very  characteristic  of 
morphine  but  requires  more  than  traces  for  a  satisfactory  result. 


Purification  of  Impure  Morphine 

When  the  chloroform  residue  is  too  impure,  especially  if  red 
or  brown,  it  must  be  purified.  Dissolve  in  hot  amyl  alcohol 
and  shake  the  solution  thoroughly  with  several  portions  of  hot 
water  containing  a  few  drops  of  dilute  sulphuric  acid.  The  acid 
dissolves  the  morphine,  whereas  the  amyl  alcohol  retains  most  of 
the  coloring  matter.  Add  ammonium  hydroxide  solution  in 
excess  to  the  acid  solution  and  extract  several  times  -u-ith  hot 
chloroform.  The  morphine  obtained  by  evaporation  of  the 
chloroform  should  be  nearly  pure. 

^  Antipyrine  and  caffeine  may  also  be  in  this  residue  (see  above). 


126  DETECTION   OF   POISONS 


MORPHINE 


Morphine,   C17H19NO3,  crystallizes  from  dilute  alcohol  in  shining  prisms 

which  are  colorless  and  transparent  and  but  slightly  soluble  in  water  (i :  5000 

at  15°;  and  1:500  at  100°),      These  solutions  are  very 

TT     TT       I     ^       bitter  and  have  an  alkaline  reaction.     Crystalline  mor- 

C      C      N  phine  is  insoluble  in  ether  and  benzene.     The  amor- 

^\X\/'\        phous  alkaloid  is  soluble  in  amyl  alcohol,  hot  chloroform 

HC      C      CH  CH2   and  acetic  ether.     Solutions  of  the  hydroxides  of  ammo- 

trn  r^  r'  r^  X-a  ^ia,  potassium  or  sodium  and  sodium  carbonate  solution 
±1U.U       L-       U      Crl2  .    .  ...  ,      .  ,  ,  . 

•^ /\    /\    /        precipitate  free  morphine  from  solutions  of  morphine 

C      C      CH       salts. 

O— c     CH2         Constitution. — Morphine    is    a    monacid, 

-^C^  tertiary  base  whose   nitrogen  is   in   union 

/\         with   three   atoms   of   carbon.      The   three 

oxygen  atoms  have  different  functions.     One 

is  a  phenolic  hydroxyl  and  gives  to  morphine  the  character  of 

a  monatomic  phenol.     Consequently  when  sodium  hydroxide 

solution  is  added  drop  by  drop  to  a  morphine  salt  solution, 

there  is  first  a  precipitate  of  crystalline  morphine  (a)  which  is 

freely  soluble  in  excess  of  alkali  (j8)  but  is  again  precipitated  on 

addition  of  ammonium  chloride  solution  (7) : 

(a)  Ci7Hi8N02(OH).HCl  +  NaOH      =  CnHisNOaCOH)     +  H2O  +  NaCl, 

(/3)    CiyHisNOaCOH)  +  NaOH      =  CuHigNOaCONa)   +  H2O, 

(7)    Ci7Hi8N02(ONa)         +  (H4N)C1  =  CnHisNOaCOH)     +  NH3  +  NaCl. 

Hydrogen  of  this  phenolic  hydroxyl  may  be  replaced  also  by 
alkyl  groups  and  acid  radicals.  In  codeine  this  hydrogen  is 
replaced  by  methyl.  A  second  oxygen  atom  of  morphine  is 
alcoholic  and  the  third  is  indifferent.  The  latter  like  the  oxygen 
of  an  ether  is  combined  with  two  carbon  atoms  and  forms  a  so- 
called  bridge-oxygen  atom. 

Of  the  17  carbon  atoms  of  morphine  14  belong  to  the  phenan- 
threne  nucleus,^  since  the  nitrogen-free  cleavage  products  of 

CH=CH                       CH=CH 
^Phenanthrene,  Ci4Hio,HC^                ^C    —     C^  ^^^'     occurs    in 

CH C  C CH 

\  / 

CH=CH 

coal-tar  together  with  anthracene.  It  forms  colorless  crystals  which  melt  at  99° 
and  boil  at  340°.  It  is  readily  soluble  in  ether  or  benzene  and  with  difi&culty  in 
alcohol.     Phenanthrene  solutions  exhibit  bluish  fluorescence. 


NON-VOLATILE   POISONS  127 

morphine  and  codeine,  namely,  morphol  and  morphenol,  have 
been  identified  as  phenanthrene  derivatives.  R.  Pschorr  has 
synthesized  morphol  which  is  3,4-dioxyphenanthrene.  Mor- 
phenol contains  two  hydrogen  atoms  less  and  may  be  converted 
into  morphol  by  reduction  with  nascent  hydrogen.  These 
two  phenanthrene  derivatives  have  the  following  structural 
formulae : 

H  H 

C  C 

/\  /\     . 

HC      CH  HC      CH 

I       II  I       II 

C      CH  C      C 

/\/  /\/\ 

HC      C  HC      C       \ 

II     I  II     I      o 

HC      C  HC      C       / 

\/\  \/\/ 

C      C.0H(4)  C      C 

I        II  I        II 

(i)HC      C.0H(3)  HC      C.OH 

\/  \/ 

(2)C  C 

H  H 

Morphol  Morphenol 

By  distillation  over  zinc  dust  morphenol  may  be  reduced  to 
phenanthrene. 

The  structural  formula  of  morphine  written  above  was  pro- 
posed by  R.  Pschorr^  and  seems  to  explain  most  satisfactorily 
the  reactions  of  this  alkaloid. 

Morphine  is  easily  oxidized.  This  may  be  brought  about  in 
alkaline  solution  by  atmospheric  oxygen.  Potassium  permanga- 
nate or  ferricyanide  and  ammoniacal  copper  solution  may  also 
be  used.  As  a  result  the  non-toxic  oxy-dimorphine,  also  called 
pseudomorphine,  which  is  soluble  in  caustic  alkali,  is  formed: 

2C17H19NO3  +    O    =    (Cl7Hi8N03)2  +  H2O. 

Morphine  Oxydimorphine 

Detection  of  Morphine 

I.  Nitric  Acid  Test. — Concentrated  nitric  acid  dissolves  mor- 
phine with  a  blood  red  color  which  gradually  changes  to  yellow. 
Stannous  chloride  or  ammonium  sulphide  solution  will  not  re- 

^  Berichte  der  Deutschen  chemischen  Gesellschaft  40,  19S4  (1907). 


128  DETECTION   OP   POISONS 

store  the  violet  color  of  a  solution  that  has  become  yellow. 
(Distinction  from  brucine.) 

2.  Husemann's  Test. — Dissolve  morphine  upon  a  watch  glass 
in  a  few  cc.  of  concentrated  sulphuric  acid.  The  solution  is  col- 
orless. Heat  for  30  minutes  upon  the  water-bath,  or  over  a 
small  flame  for  a  very  short  time  until  white  fumes  arise.  A 
reddish  or  brownish  color  appears.  Cool  and  add  1-2  drops  of 
concentrated  nitric  acid.  A  fugitive,  reddish  violet  color  ap- 
pears and  soon  changes  to  blood  red  or  yellowish  red.  This 
color  gradually  disappears. 

A  preferable  procedure  is  to  dissolve  morphine  in  cold  con- 
centrated sulphuric  acid  and  add  a  trace  of  concentrated  nitric 
acid  after  the  solution  has  stood  in  a  desiccator  24  hours.  A 
small  crystal  of  potassium  nitrate  or  chlorate  may  be  substituted 
for  nitric  acid. 

Frequently  impure  morphine  is  obtained  from  the  chloroform  extract  of  a 
solution  prepared  from  animal  material.  Such  a  residue  gives  a  more  or  less 
highly  colored  solution  with  sulphuric  acid.  Heat  usually  intensifies  the  color. 
But  even  under  these  conditions  it  is  possible  to  detect  the  red  color  caused  by 
nitric  acid  or  potassium  nitrate. 

3.  Pellagri's  Test. — Proceed  as  described  for  codeine.  (See 
page  108.)  Avoid  excess  of  alcoholic  iodine  solution,  otherwise 
the  latter  may  mask  the  green  color. 

4.  Froehde's  Test. — This  reagent  dissolves  morphine  with  a 
violet  color  which  passes  through  blue  to  dirty  green  and  finally 
to  faint  red.     These  colors  vanish  on  addition  of  water. 

5.  Formaldehyde -Sulphuric  Acid  Test. — The  solution  used 
for  this  test  is  called  Marquis'  reagent^  With  a  trace  of  mor- 
phine it  produces  a  purple-red  color  which  changes  to  violet 
and  finally  becomes  pure  blue.  This  blue  solution,  kept  in  a 
test-tube  and  only  slightly  exposed  to  air,  retains  its  color  for 
some  time.  Codeine  and  apomorphine  give  the  same  violet 
color.  Narcotine  also  gives  violet  solutions  but  they  become 
olive  green  and  finally  yellow.  Oxy-dimorphine  gives  a  green 
color. 

^  Mix  2-3  drops  of  40  per  cent,  formaldehyde  solution  with  5  cc.  of  concen- 
trated sulphuric  acid  and  use  a  few  drops  of  this  mixture  for  the  morphine  test. 


NON-VOLATILE    POISONS  129 

6.  Iodic  Acid  Test. — Shake  a  solution  of  morphine  in  dilute 
sulphuric  acid  with  a  few  drops  of  iodic  acid  and  chlorofcjrm. 
Morphine  will  liberate  iodine  which  will  dissolve  in  chloroform 
with  a  violet  color. 

Obviously  this  delicate  test  is  conclusive  for  morphine  only  in  the  absence  of 
other  reducing  substances. 

7.  Ferric  Chloride  Test. — Add  1-2  drops  of  neutral  ferric 
chloride  solution  to  a  neutral  solution  of  a  morphine  salt.  A 
blue  color  appears.  In  testing  the  chloroform  residue,  dissolve 
in  a  little  very  dilute  hydrochloric  acid.  Evaporate  this  solu- 
tion to  dryness  upon  the  water-bath,  dissolve  the  residue  in  pure 
water  and  add  a  drop  of  ferric  chloride  solution. 

8.  Lloyd's  Test. — ^Lloyd  has  found  that  a  mixture  of  morphine, 
hydrastine  and  concentrated  sulphuric  acid  alone  without 
potassium  dichromate  will  produce  the  same  violet  color  given 
by  the  latter  with  a  solution  of  strychnine  in  concentrated  sul- 
phuric acid.  Lloyd's  reaction  is  of  value  in  the  detection  of  mor- 
phine or  hydrastine  only  when  more  than  traces  of  both  alka- 
loids are  present.  A.  Wangerin^  considers  these  reactions 
characteristic  only  when  0.005-0.01  gram  of  morphine  and 
0.002-0.01  gram  of  hydrastine  are  present. 

Make  an  intimate  mixture  of  about  these  quantities  of  both 
alkaloids  upon  a  watch  glass.  Add  5  drops  of  pure  concen- 
trated sulphuric  acid  and  stir  the  mixture  for  10  minutes 
over  a  white  background.  In  the  center  the  color-tone  is  a 
clear  red-violet  and  more  or  less  of  a  blue-violet  in  the  thinner 
marginal  region.  Apomorphine  hydrochloride,  treated  in  the 
same  way  with  hydrastine  and  concentrated  sulphuric  acid, 
gives  almost  the  same  reaction  as  morphine. 

9.  Prussian  Blue  Test. — Add  a  few  drops  of  a  dilute  mixture 
of  ferric  chloride  and  potassium  ferricyanide  solutions  to  a 
morphine  salt  solution.  A  deep  blue  color  appears.  Consid- 
erable morphine  produces  a  precipitate  of  Prussian  blue. 

Potassium  ferricyanide  oxidizes  morphine  to  oxy-dimorphine: 
2C17H19NO3  +  2KOH  +  K6Fe2(CN)i2  =  2H2O  -1-  (Ci7Hi8X03)2  +  2K4Fe(CX)6 

Morphine  Potassium  Oxy-dimorphine  Potassium 

ferricyanide  ferrocyanide 

1  Pharmazeutische  Zeitung  46,  57(1903). 
9 


130  DETECTION   OE   POISONS 

Potassium  ferrocyanide  then  forms  Prussian  blue  with  ferric 
chloride. 

10.  Silver  Test. — Warm  a  morphine  salt  solution  with  silver 
nitrate  and  excess  of  ammonium  hydroxide  solution.  Mor- 
phine produces  a  gray  precipitate  of  metallic  silver. 

11.  Bismuth  Test. — Dissolve  morphine  in  concentrated 
sulphuric  acid  and  sprinkle  a  little  bismuth  subnitrate  on  the 
surface  of  the  solution.     A  dark  brown  color  appears. 

12.  G.  Fleury's  Test.^ — Dissolve  morphine  in  a  little  very 
dilute  sulphuric  acid  (about  0.05  normal),  add  some  lead  di- 
oxide (Pb02)  and  shake  for  6-8  minutes.  A  pale  rose  color 
appears.  Addition  to  the  filtrate  of  ammonium  hydroxide 
solution  in  excess  produces  a  brown  color  which  persists  for 
several  hours.  When  the  quantity  of  substance  is  very  small, 
stir  on  a  porcelain  color  plate  for  6-8  minutes  with  a  drop  of 
dilute  sulphuric  acid  and  a  minute  particle  of  lead  dioxide. 
When  the  insoluble  matter  has  settled,  tilt  the  porcelain  plate 
so  that  the  clear  solution  runs  up  the  side.  A  drop  of  ammonium 
hydroxide  solution  now  gives  a  brown  color. 

13.  Dan  Radulescu's  Test.^ — Add  a  small  particle  of  sodium 
nitrite  to  a  very  dilute  morphine  salt  solution,  then  a  dilute 
acid  and  render  alkaline  with  concentrated  potassium  hydroxide 
solution  before  all  the  gas  has  escaped.  The  solution  when  con- 
centrated has  a  pale  rose  to  a  deep  ruby  red  color.  Acids  dis- 
charge but  alkalies  restore  this  color.  This  reaction  is  said  to 
be  characteristic  of  morphine  bases  and  especially  adapted 
for  the  detection  of  morphine  in  mixtures. 

General  Alkaloidal  Reagents. — The  reagents  of  this  class 

especially  sensitive  toward  solutions  of  morphine  salts  are: 

lodo-potassium  iodide  Potassium  bismuthous  iodide 

Phospho-tungstic  acid  Phospho-molybdic  acid 

Potassium  mercuric  iodide  Gold  chloride. 

Plantinic  chloride  after  some  time  causes  a  granular  orange- 
yellow  precipitate.  Tannic  acid  causes  no  precipitate,  or  at 
most  only  a  very  slight  cloudiness  which  becomes  somewhat 
more  pronounced  with  time. 

^Annales  de  Chimie  analytique  appliquee  6,  417  (1907). 
'  Chemisclies  Zentralblatt  1906,  i,  1378. 


NON-VOLA'IILK    POISONS  1  •'>  I 

Behavior  of  Morphine  in  the  Animal  Organism. — The  mucous  lining  of  the 
stomach,  rectum  or  respiratory  passages  as  well  as  open  wounds  absorb  mor- 
phine. The  alkaloid  injected  hypodermically  acts  more  rapidly  and  more  po- 
tently than  when  absorbed  from  the  stomach.  Marquis'  found  that  morphine 
disappears  very  quickly  from  the  blood  but  is  firmly  retained  by  certain  organs 
like  the  brain.  Some  absorbed  morphine  is  conjugated  with  glycuronic  acid  and 
some  is  oxidized  but  the  rest  of  the  alkaloid  is  eliminated  unchanged.  Faust 
has  found  that  morphine  is  transformed  or  destroyed  only  in  men  and  animals 
habituated  to  the  poison  but  is  eliminated  unchanged  nearly  quantitatively  in 
the  faeces  in  the  case  of  organisms  not  immunized.  Morphine  appears  in  the  urine 
only  in  very  small  quantity  after  medicinal  doses.  In  men  and  dogs  a  not  insig- 
nificant quantity  of  the  morphine  taken  is  eliminated  by  the  glands  of  the  gastro- 
intestinal tract,  even  when  the  alkaloid  has  been  subcutaneously  injected. 

Marquis  found  that  more  than  30  per  cent,  of  intravenously  injected 
morphine  is  deposited  in  the  liver  in  the  course  of  15  minutes.  The  alkaloid  is 
present  at  first  in  this  organ  in  the  free  state  and  then  is  soon  combined  or  trans- 
formed. The  conjugation  of  morphine  in  the  brain  also  begins  very  soon.  Free 
morphine  is  also  rapidly  changed  in  the  blood,  spleen,  kidneys  and  in  the  mucous 
lining  of  the  intestines.  Marquis  states  that  always  in  acute  and  even  more  so 
in  chronic  morphine  poisoning  a  large  quantity  of  the  poison  leaves  the  blood  and 
is  stored  in  the  salivary  glands,  mucous  lining  of  the  stomach  and  large  intestine, 
kidneys,  spleen,  liver  and  is  withdrawn  by  these  organs  from  the  brain  and  spinal 
cord. 

Morphine  is  quite  resistant  to  putrefaction.  The  author^  detected  this  alka- 
loid positively  in  animal  material  containing  morphine  (stomach  and  intestines 
together  with  contents)  which  had  stood  for  15  months  in  a  glass  vessel  and  had 
completely  putrefied  in  presence  of  insufficient  air. 

NARCEINE 

OCH3  Narceine,   C23H27NO3.3H2O,   crystallizes    from 

C  water  or  alcohol  in  prisms  which  melt  at  165° 

^\  when  air  dried.     The  alkaloid  has  a  faintly  bitter 

^9      C.OCH3  j^g(.g_     Though  only  slightly  soluble  in  cold  water, 

HC      (j.COOH  ^^  ^^  freely  soluble  in  hot.     When  a  hot  saturated 

^/Z  aqueous  solution  of  narceine  is  cooled,  it  solidifies 

C  to  a  crystalline  mass.     Narceine  is  insoluble  in 

X^  ether,  benzene  or  petroleum  ether  and  is  soluble 

CjjgO.       I  °^ly  "^ith  difficulty  in  cold  alcohol,  amyl  alcohol 

C      CH2  or  chloroform.     In  detecting  narceine  it  is  im- 

v\/         /CH3       portant  to  know  that  it  is  not  extracted  bv  ether, 
O  C      C       N<  " 

TT  p  /    ■  I        II        I  \pTT       benzene  or  petroleum  ether  from  a  solution  ren- 

^O.C      C      CH2  dered  alkaline  by  potassium  or  sodium  hydroxide 

\/\/  solution.     It  is,  however,  extracted  by  hot  chloro- 

C      C  form  or  amyl  alcohol  from  an  aqueous  solution  ren- 

^  dered  alkaline  by  ammonium  hydroxide  solution. 

'Arbeiten  des  Dorpater  Instituts,  ed  Kobert,  14  (1896). 

'Berichte  der  Deutschen  Pharmazeutischen  Gessellschaft  11,  494  (1901). 


132 


DETECTION   OF   POISONS 


Constitution. — Narceine  is  a  weak  tertiary  base  in  which  two 
methyl  groups  are  attached  to  nitrogen.  By  means  of  Zeisel's 
method  it  may  be  shown  that  the  molecule  also  contains  three 
methoxyl  groups.  Narceine,  being  soluble  in  caustic  alkalies 
and  forming  esters  with  alcohols,  must  contain  a  carboxyl 
group.  The  alkaloid  must  also  contain  a  carbonyl  group  (CO), 
since  it  forms  a  hydrazone  with  phenyl-hydrazine.  The  nar- 
ceine formula  above  may  therefore  be  resolved  into: 

C23H27N08=  CieHiiON  (CH3)2  (OCH3)3  (CO)  (COOH). 

The  narceine  molecule  contains  neither  an  alcoholic  nor  a 
phenolic  hydroxyl  group,  since  it  forms  no  acetyl  derivative 
with  acetic  anhydride.  There  is  a  close  relationship  between 
narceine  and  narcotine.  By  heating  narcotine  iodo-methylate 
with  sodium  hydroxide  solution  Roser  converted  this  compound 
into  a  base  called  pseudo-narceine.  Freund  has  recently  shown 
that  Roser's  pseudo-narceine  is  identical  with  the  opium  alka- 
loid narceine  and  explains  the  conversion  of  narcotine  into  nar- 
ceine by  saying  that  the  iodo-methylate  loses  i  molecule  of 
hydriodic  acid  and  takes  up  i  molecule  of  water: 


OCH3 


OCH3 


H2C 


c 

C 

/\ 

/\ 

HC      C.OCH3 

HC       C.OCH3 

1         II 
HC       C.COOH 

HC      C.CO 

\/ 

\/ 

c 

+  H2O 

c 

1 

CHsOJHiC— 0 

-  HI 

CH3O      CO 

C  HC 

C      CH2 

/\/\  /ilj 

/-\/    /CH3 

/O.C      C     N< 

H2C<       1       il        1  \CH3 

^O.C     C      CH2 

/O.C      C      N— CH3 
C<      1        II        |\CH3 
^O.C      C      CH2 

\/\/ 

\/\/ 

c    c 

c    c 

H    H2 

H     H2 

Narcotine  iodo-meth 

ylate 

Narceine 

All  the  reactions  and  transformations  of  narceine  can  easily 
be  explained  on  the  basis  of  this  structural  formula. 


NON-VOLATILE    POISONS  133 

Detection  of  Narceine 

1.  Sulphuric  Acid  Test. — Concentrated  sulphuric  acid  dis- 
solves narceine  with  a  grayish  brown  color,  which  gradually 
changes  to  blood  red.  This  reaction  takes  place  at  once  with 
heat. 

2.  Dilute  Sulphuric  Acid  Test. — Narceine,  warmed  in  a  porce- 
lain dish  upon  the  water-bath  with  dilute  sulphuric  acid  until 
a  certain  concentration  is  reached,  gives  rise  to  a  fine  violet 
color  which  changes  after  long  heating  to  cherry  red. 

3.  Froehde's  Test. — At  first  a  solution  of  narceine  in  this 
reagent  has  a  brownish  green  color  which  gradually  changes 
to  green  and  finally  to  red.     Gentle  heat  hastens  this  reaction. 

4.  Iodine  Test. — Aqueous  iodine  solution  (iodine  water)  or 
iodine  vapor  colors  solid  narceine  blue. 

Morphine  interferes  with  or  entirely  prevents  this  reaction. 

5.  Erdmann's  Test. — This  reagent,  as  well  as  concentrated 
nitric  acid,  dissolves  narceine  with  a  yellow  color  which  heat 
changes  to  dark  orange. 

6.  Chlorine -Ammonia  Test.^ — Pour  a  few  drops  of  chlorine 
water  upon  narceine  and  add,  while  stirring,  a  few  drops  of 
ammonium  hydroxide  solution.  A  deep  red  color  immediately 
appears. 

7.  Resorcinol-Sulphuric  Acid  Test.^ — Mix  thoroughly  upon 
a  watch  glass  resorcinol  (o.oi  to  0.02  gram)  with  10  drops  of 
pure  concentrated  sulphuric  acid.  Add  a  trace  of  narceine 
(about  0.002  to  0.005  gram)  and,  while  stirring,  warm  the  in- 
tensely yellow  solution  upon  a  boiling  water-bath.  A  carmine 
red  to  cherry  red  color  appears.  As  the  solution  cools,  this 
color  begins  at  the  margin  to  change  gradually  to  more  of  a 
blood  red  and  finally  after  several  hours  to  orange-yellow. 

8.  Tannin-Sulphuric  Acid  Test.— Mix  narceine  (0.002  to 
o.oi  gram)  with  tannin  (o.oi  to  0.02  gram)  and  10  drops  of 
pure  concentrated  sulphuric  acid.  Heat  with  constant  stirring 
upon  the  water-bath  and  the  color  of  the  solution,  which  is 
yellowish  brow-n  at  first,  soon  becomes  pure  green.     If  heat  is 

1  A.  Wangerin,  Pharmaceutische  Zeitung,  47,  916  (1902). 


134  DETECTION   OF   POISONS 

applied  for  some  time,  the  green  color  changes  to  blue-green 
and  finally  through  a  more  or  less  blue  tone  to  a  dirty  green. 

Tannin-sulphuric  acid  gives  a  similar  color  test  with  narcotine  and  hydras- 
tine  which  closely  resemble  narceine  in  constitution. 

Of  the  general  alkaloidal  reagents  potassium  zinc  iodide^ 
precipitates  narceine  even  in  a  dilution  of  i  :  looo.  It  is  a 
white,  filiform  precipitate  which  after  a  time  becomes  blue. 
This  blue  color  appears  immediately,  if  a  trace  of  iodine  solu- 
tion is  added  to  the  reagent. 

Of  the  other  general  reagents  iodo-potassium  iodide,  potassium  mercuric 
iodide,  potassium  bismuthous  iodide  and  phospho-molybdic  acid  are  characterized 
by  considerable  delicacy  toward  narceine. 

SYNOPSIS  OF  GROUP  II 

Stas-Otto  Method 

A.  Ether  Extract  of  Acid  Solution  may  Contain : 

Picrotoxin. — Very  bitter.  Reduces  Fehhng's  solution  with 
heat. 

Melzer's  test:  red  streaks  radiating  from  picrotoxin  with 
alcohoKc  benzaldehyde  +  cone.  H2SO4. 

Cone.  H2SO4:  soluble  with  yellow  or  orange-red  color;  drop 
of  K2Cr207  +  Aq  has  brown  margin. 

Langley's  test:  picrotoxin  +  3  parts  KNO3,  moistened  with 
cone.  H2SO4,  red  with  excess  of  saturated  NaOH  +  Aq. 

Colchicin. — Very  bitter.  Yellowish  and  amorphous.  Dilute 
mineral  acids  render  aqueous  solutions  intensely  yellow. 

Cone.  HNO3:  soluble  with  dirty  violet  color  changing  to 
brownish  red  and  finally  to  yellow;  excess  of  KOH  -f-  Aq 
renders  orange-yellow  or  orange-red. 

ZeiseFs  test:  boil  yellow  colchicin  solution  in  cone.  HCl  in  test- 
tube  2-3  minutes  with  2  drops  of  FeClsH-Aq.  Green  or  olive- 
green  when  cold,  especially  if  diluted  with  equal  volume  of  water. 

Picric  Acid. — Very  bitter.  Yellow.  Material  and  extracts 
more  or  less  intensely  yellow. 

^  See  page  312  for  the  preparation  of  this  reagent. 


NON-VOLATILE   POISONS  135 

Isopurpuric  acid  test:  aqueous  picric  acid,  gently  warmed 
with  a  few  drops  of  saturated  KCN  +  Aq,  gives  red  color. 

Picraminic  acid  test:  aqueous  picric  acid,  warmed  with  few 
drops  of  (H4N)2S  +  Aq,  becomes  red. 

Dyeing  test:  aqueous  picric  acid  dyes  wool  and  silk  intense 
yellow  but  not  cotton. 

Acetanilide. — Faint,  burning  taste. 

Indophenol  test:  heat  with  a  few  cc.  of  cone.  HCl  and  evap- 
orate to  about  20  drops.  Cool,  add  aqueous  phenol  solution 
and  then  calcium  hypochlorite  solution  drop  by  drop.  Mix- 
ture, shaken  with  excess  of  ammonia,  becomes  dirty  red  to 
blue-violet  and  blue. 

PhenyKsocyanide  test:  boil  with  KOH  +  Aq  and  then  add 
a  little  chloroform.     Odor  of  phenylisocyanide. 

Isolation  of  aniline:  boil  several  minutes  with  alcoholic  KOH, 
dilute  with  water  and  extract  with  ether.  Evaporation  of 
solvent  leaves  oily  drops  of  aniline.  Dissolve  in  water  and  test 
with  calcium  hypochlorite. 

Phenacetine. — Tasteless.  Gives  indophenol  but  not  phenyl- 
isocyanide test. 

Cone.  HNO3:  yellow  color  even  cold.  Dil.  HNO3  dissolves 
with  yellow  or  orange-yellow  color,  if  heated.  Yellow  nitro- 
phenacetine  crystalhzes  as  saturated  solution  cools. 

Salicylic  Acid. — Sweet,  acidulous,  harsh  taste. 

FeCls  -f-  Aq:  aqueous  solutions  colored  blue-violet;  if  dilute, 
more  of  a  red-violet. 

Millon's  test:  red  color  upon  warming. 

Br2  -h  Aq:  yellowish  white,  crystalline  precipitate. 

Veronal. — Bitter.     CrystalHne. 

Dissolve  ether  residue  in  very  little  NaOH  +  Aq  or  (H4N")- 
OH  -\-  Aq,  filter  and  acidify  filtrate  with  dil.  HCl.  Veronal 
crystallizes.  Wash  with  a  little  cold  water,  dry  and  determine 
melting  point  (187-188°).  The  crystals  mLxed  with  pure 
veronal  should  have  same  melting  point. 

Antipyrine.— Mild,  bitter  taste.  Examine  aqueous  solution 
of  ether  residue  for  antipyrine. 

FeCls  +  Aq:  red  color. 


136  DETECTION    OF   POISONS 

HNO3:  green  color  with  1-2  drops  of  fuming  acid.  Heat 
and  a  few  more  drops  of  fuming  acid  change  green  color  to  red. 

Most  of  the  antipyrine  in  ether  extract  of  alkaline  solution 
(seeB). 

Caffeine. — Faintly  bitter. 

CI2  +  Aq:  evaporated  upon  water-bath  with  saturated 
CI2  +  Aq,  gives  red-brown  residue  which  turns  purplish  red 
moistened  with  very  little  (H4N)0H  +  Aq. 

Most  of  the  caffeine  in  ether  extract  of  alkaline  solution 
(see  B). 

Cantharidin.^^ — Rhombic  leaflets  from  ether  solution. 

Physiological  test:  triturate  residue  with  few  drops  of  almond 
oil  and  test  mixture  as  vesicant  by  applying  to  upper  part  of 
the  arm. 

B.  Ether  Extract  of  Alkaline  Solution  may  Contain : 

Coniine. — Yellow  oil  drops  with  penetrating  odor. 

Cold  saturated  aqueous  solution  becomes  milky  when  warmed. 

Spontaneous  evaporation  with  a  drop  of  HCl  gives  coniine 
hydrochloride  as  doubly  refractive  crystals  which  are  needle 
or  prism  shaped  and  sometimes  in  star-like  clusters. 

Physiological  test:  paralysis  of  peripheral  nerves. 

Nicotine. — ^Liquid.  Remains  dissolved  in  residual  water 
upon  evaporation  of  ether  and  has  faint  tobacco  odor. 

Melzer's  test:  red  color,  heated  with  2-3  cc.  of  epichlorohy- 
drin. 

Schindelmeiser's  test:  nicotine,  after  standing  several  hours 
with  a  drop  of  formaldehyde  solution,  gives  an  intense  red  color 
with  a  drop  of  cone.  HNO3. 

Roussin's  test:  ether  solution  of  iodine  after  some  time  pro- 
duces ruby  red,  crystalline  needles. 

Aniline.^ — Yellow,  reddish  or  brownish  oil  drops  from  evapora- 
tion of  ether  extract.  (See  page  56.  "Synopsis  of  Group  I" 
for  further  details.) 

^  Cantharidin  is  taken  up  in  Chapter  IV  of  this  book  upon  page  196.  Ether 
extracts  this  compound  from  acid  solution  but  it  dissolves  with  difficulty  in  this 
solvent  (o.ii  :  100  at  18°). 


NON-VOLATILE   POLSONS  137 

Veratrine. — Cone.  H2SO4 :  soluble  with  yellow  color,  gradually 
changing  to  orange,  then  to  red  and  finally  to  cherry  red.  Gen- 
tle heat  hastens  these  changes.  Solution  at  first  shows  greenish 
yellow  fluorescence. 

Froehde:  same  color  changes 'as  with  cone.  H2SO4. 

Cone.  HCl:  very  stable  red  color  when  heated  in  test-tube 
upon  water-bath. 

Weppen's  test:  mixed  with  6  times  the  quantity  of  cane 
sugar  +  a  few  drops  of  cone.  H2SO4,  gradually  becomes  green 
and  finally  blue.  Cone.  H2SO4  containing  furfurol  may  be 
used  instead. 

Vitali's  test:  same  as  for  atropine  (see  below). 

Strychnine.^ — Fine,  crystalline  needles  having  a  very  bitter 
taste  upon  evaporation  of  ether  extract. 

Oxidation  test:  colorless  solution  in  cone.  H2SO4  becomes 
evanescent  blue  or  blue-violet  with  a  little  solid  K2Cr207. 
Same  color  given  by  Mandelin's  reagent  but  more  permanent. 

Brucine. — Cone.  HNO3:  dissolves  with  blood  red  color  soon 
changing  to  reddish  yellow  and  yellow.  Dilution  of  yellow 
solution  in  a  test-tube  with  a  little  water  and  addition  drop  by 
drop  of  dilute  SnCl2  +  Aq  changes  yellow  to  violet. 

Careful  addition  of  solution  in  dil.  HNO3  to  cone.  H2SO4  as 
upper  layer  produces  red  or  yellowish  red  zone. 

Atropine. — Vitali's  test:  evaporated  upon  water-bath  in  por- 
celain dish  with  a  little  fuming  HNO3,  gives  yellowish  residue 
which  becomes  violet  when  moistened  with  alcoholic  KOH. 
Hyoscyamine  and  scopolamine  also  give  this  test.  Strychnine 
and  veratrine  behave  similarly. 

Physiological  test:  enlargement  of  pupil  of  eye  caused  by  a 
single  drop  of  solution  i  :  130,000. 

Cocaine. — Free  base,  precipitated  by  KOH  +  Aq  from  not 
too  dilute  cocaine  salt  solution,  forms  oil  drops  soon  becoming 
solid  and  crystalline. 

Benzoyl  group:  heat  5  minutes  in  a  test-tube  upon  boiling 
water-bath  with  i  cc.  cone.  H2SO4.  Odor  of  methyl  benzoate 
upon  addition  of  2  cc.  of  water.     Upon  cooling,  benzoic  acid 


138  DETECTION   OF  POISONS 

separates.  This  acid,  washed  and  dried,  recognized  by  melting 
point  (120°)  and  by  tendency  to  sublime. 

Physiological  test:  anesthesia  of  the  tongue. 

Codeine. — Cone.  H2SO4:  soluble  without  color.  Reddish  or 
more  bluish  upon  long  standing,  or  at  once  upon  gentle  warming. 

Oxidation:  Deep  blue  or  blue-violet,  when  warmed  with 
cone.  H2SO4  and  KH2ASO4,  or  with  a  little  FeCls+Aq. 

Froehde:  yellowish  color  soon  changing  to  green  and  to  blue 
upon  gentle  warming. 

Sugar  test:  purple-red  color  upon  gently  warming  with  cone. 
H2SO4  and  a  little  cane  sugar.     Due  to  furfurol  formed. 

Formahn  test:  dissolves  in  cone.  H2SO4  containing  formalde- 
hyde with  reddish  violet  color  soon  changing  to  permanent  blue- 
violet. 

Pellagri's  test:  given  by  codeine  (see  apomorphine,  page  140). 

Hydrastine. — Froehde:  dissolves  with  fairly  permanent  green 
color  later  changing  to  brown. 

Mandelin:  dissolves  with  reddish  color  gradually  changing 
to  orange-red. 

Fluorescence:  intense  blue  fluorescence  (characteristic)  upon 
shaking  dil.  H2SO4  solution  with  very  dilute  KMn04  +  Aq  added 
carefully  drop  by  drop. 

Quinine. — Amorphous  varnish  having  very  bitter  taste  from 
ether. 

Fluorescence:  blue  fluorescence  in  dil.  H2SO4. 

Thalleioquin  test :  emerald  green  color,  upon  adding  i  cc.  satu- 
rated CI2  +  Aq  to  dilute  acetic  acid  solution  and  then  at  once 
excess  of  (H4N)0H-|-Aq  drop  by  drop. 

Herapathite  test:  heat  to  boiling  with  10  drops  of  mixture 
(30  drops  acetic  acid  4-20  drops  absolute  alcohol  +  i  drop  dil. 
H2SO4)  and  add  i  drop  alcoholic  iodine  solution  (1:10). 
Shining,  olive  green  leaflets,  appearing  cantharides  green  by 
reflected  light. 

Antipyrine. — Freely  soluble  in  water.  Neutral.  Mildly 
bitter. 

Dissolve  ether  residue  in  little  water  and  test  for  antipyrine  as 
directed  in  A  (see  page  135). 


NON-VOLATILE   POISONS  139 

Pyramidone. — Fine  needles  from  ether.  Freely  soluble  in 
water.     Neutral. 

FeCls  +  Aq:  aqueous  solution  blue-violet  or  more  red- violet. 

HNO3:  fuming  acid  renders  aqueous  solution  blue  to  blue- 
violet. 

Caffeine. — Concentric  clusters  of  shining  needles  from  ether. 
Mild,  bitter  taste.     Fairly  soluble  in  water.     Neutral. 

Apply  tests  described  under  A  (see  page  136). 

Physostigmine. — (H4N)0H-|- Aq:  evaporated  with  (H4N)- 
OH  H-  Aq,  gives  blue  residue  soluble  in  alcohol  with  same  color. 

Physiological  test:  causes  contraction  of  pupil  of  eye. 

Narcotine. — Not  bitter.     Neutral. 

Froehde:  soluble  with  green  color.  A  concentrated  reagent 
(0.05  gram  (H4N)2Mo04  to  i  cc.  cone.  H2SO4)  gives  greenish 
color  at  first  which  gradually  changes  to  cherry  red  and  to  a 
blue  from  margin  toward  center. 

Erdmann:  soluble  with  fine  red  color. 

Papaverine. — Tasteless,  colorless,  neutal  prisms. 

Cone.  H2SO4:  pure  alkaloid  soluble  without  color.  Heat 
produces  dark  violet  color. 

Froehde:  soluble  with  green  color,  soon  changing  when  warmed 
to  blue,  violet  and  finally  cherry  red. 

HNO3-H2SO4  test:  cone.  H2SO4  containing  HNO3,  or  cone. 
HNO3  itself,  gives  a  dark  red  solution. 

Thebaine. — Tasteless,  colorless,  alkahne  prisms. 

Cone.  H2SO4:  soluble  with  deep  red  color.  Froehde  and 
Erdmann  behave  similarly. 

C.  Ether  Extract^  of  Ammonia  Solution  may  Contain : 

Apomorphine. — Residue  amorphous  and  usually  green. 
H2SO4-HNO3  test:  solution  in  cone.  H2SO4  colored  evanescent 
violet,  then  reddish  yellow  or  orange  by  drop  of  cone.  HNO3. 
Froehde:  soluble  with  green  or  violet  color. 
Pellagri's  test:  dissolve  in  dil.  HCl,  add  excess  of  XaHCOs, 

^  Unless  the  tartaric  acid  and  alkaline  solntions,  as  well  as  their  ether  extracts, 
behave  as  described  on  page  121,  that  is  to  saj',  have  a  green  or  red  color, 
omit  this  extraction. 


140  DETECTION    OF   POISONS 

shake  well  and  add  2  drops  alcoholic  iodine  solution.     Blue  or 
emerald-green  color  soluble  in  ether  with  violet  color. 

Wangerin's  test:  1-2  drops  K2Cr207  +  Aq  (0.3  per  cent.), 
added  to  apomorphine  hydrochloride  solution,  gradually  pro- 
duces dark  green  color.  Chloroform  added  becomes  violet. 
Addition  of  dil.  SnCl2  +  Aq  produces  pure  indigo-blue  color. 

D.  Chloroform  Extract  of  Ammonia  Solution  may  Contain : 

Morphine. — Very  bitter.  Usually  amorphous.  Rarely  crys- 
talline. 

Froehde:  soluble  with  violet  color  gradually  changing  to 
dirty  green  and  finally  to  pale  red. 

Formaldehyde-H2S04:  soluble  with  purple-red  color  later  be- 
coming blue-violet  and  almost  pure  blue. 

Husemann's  test:  dissolve  in  cone.  H2SO4,  heat  over  very  small 
flame  until  abundant  white  fumes  appear,  cool  and  add  i  drop 
cone.  HNO3.  Very  evanescent,  red-violet  color  which  soon 
changes  to  blood-red  or  reddish  yellow. 

Pellagri's  test:  see  apomorphine. 

FeCls+Aq:  dissolve  in  few  drops  very  dilute  HCl,  evaporate 
to  dryness  upon  water-bath,  dissolve  in  little  water  and  add 
drop  FeCls+Aq.     Blue  color. 

Bismuth  test:  dissolve  in  cone.  H2SO4  and  sprinkle  bismuth 
subnitrate  on  surface  of  solution.     Dark  brown  color. 

Antip3rrine  and  Caffeine.-^Being  soluble  in  ether  with  some 
difficulty,  but  readily  soluble  in  chloroform,  these  substances 
may  appear  in  the  residue  from  D,  if  they  have  not  been  pre- 
viously completely  extracted  with  ether. 

Narceine. — 12  test:  blue  color  with  12  + Aq. 

Resorcinol-H2S04  test:  dissolves  in  resorcinol-H2S04,  giving 
intense  yellow  solution  which  becomes  carmine-red  or  cherry- 
red,  if  warmed  upon  the  water-bath  and  stirred. 

Tannin-H2S04  test:  dissolves  in  tannin-H2S04,  giving  yellow- 
ish brown  solution  which  becomes  pure  green,  if  warmed  upon 
the  water-bath. 


CHAPTER  III 

METALLIC     POISONS 

Destruction  of  Organic  Matter 

The  analyst  cannot  rely  upon  tests  for  poisonous  metals,  if 
animal  or  vegetable  matter  is  present.  Consequently  complete 
destruction  of  interfering  organic  substances  is  absolutely 
essential  to  success.  Description  of  a  few  of  the  more  im- 
portant methods  used  for  this  purpose  will  sufhce. 

I.  Fresenius-v.  Babo  Method^ 

The  residue  left  after  removal  of  volatile  poisons  by  steam 
distillation  may  be  used  in  this  part  of  the  analysis,  as  it  must 
contain  poisonous  metals  if  any  are  present. 

A  portion  of  the  original  material,  ^  previously  finely  chopped 
and  well  mixed  in  a  large  flask  with  enough  water  to  produce  a 
fluid  mass,  may  also  be  used.  According  to  the  quantity  of 
material,  add  lo,  20  or  30  cc.  of  pure  concentrated  hydrochloric 
acid.^  Finally  add  1-2  grams  of  potassium  chlorate,  shake  well 
and  set  the  flask  upon  a  boihng  water-bath.  Nascent  chlorine 
should  come  into  contact  with  the  material  as  intimately  as 
possible. 

When  the  mixture  is  hot  enough,  add  0.3-0.5  gram  of  potas- 
sium chlorate  at  5  minute  intervals  and  shake  the  flask  fre- 
quently. Continue  in  this  manner,  until  most  of  the  organic 
matter  is  dissolved  and  the  solution  is  pale  yellow.     Further 

1  Annalen  der  Chemie  und  Pharmazie  49,  306  (1844). 

2  Cadaveric  material  should  be  divided  as  finely  as  possible,  then  brought  to  a 
thin  mixture  by  stirring  with  12.5  per  cent.,  arsenic-free  hydrochloric  acid  and 
heated  with  frequent  shaking  \vith  1-2  grams  of  potassium  chlorate  as  directed 
above.  If  the  material  is  heated  on  the  water-bath  in  a  porcelain  dish,  it  should 
be  stirred  constantly. 

2  In  laboratory  experiments  5-10  cc.  cone,  hydrochloric  acid  is  usually  suffi- 
cient.    A  large  excess  of  hydrochloric  acid  should  be  avoided. 

141 


142 


DETECTION   OF   POISONS 


addition  of  potassium  chlorate  and  longer  heating  should 
produce  no  real  change.  Fat  especially  resists  the  action  of 
chlorine. 

When  organic  matter  is  completely  destroyed,  dilute  with 
hot  water,  adding  a  few  drops  of  dilute  sulphuric  acid  to  pre- 

cipitate  possible  barium, 
shake  and  pour  the  liquid 
through  a  wetted  filter.  If 
the  excess  of  free  hydro- 
chloric acid  is  not  too  large, 
saturate  the  filtrate  direct 
with  hydrogen  sulphide  as 
directed  on  page  145.  Other- 
wise, evaporate  the  solution 
in  a  porcelain  dish  upon  the 
water-bath  nearly  to  dryness 
to  remove  most  of  the  free 
hydrochloric  acid.  This  step 
frequently  gives  rise  to  a  dark 
brown  color  which  a  few 
crystals  of  potassium  chlorate 
will  discharge.  In  testing  for 
lead,  cadmium  and  copper,  it 
is  advisable  to  evaporate, 
because  hydrogen  sulphide 
precipitates  the  first  two 
metals  incompletely,  or  not 
at  all,  from  solutions  contain- 
ing too  much  hydrochloric 
acid. 
An  alternative  procedure  consists  in  removing  part  of  the 
free  hydrochloric  acid  from  the  filtrate,  obtained  after  treat- 
ment with  hydrochloric  acid  and  potassium  chlorate,  by  first 
evaporating  to  smaller  volume  and  then  adding  ammonium 
hydroxide  solution  until  alkaline.  Add  dilute  nitric  acid  until 
the  solution  is  faintly  acid  and  saturate  with  hydrogen  sulphide 
(seepage  145). 


Ftg.  12. 


MKIALLIC   POISONS  143 

The  residue  upon  the  filter  may  contain  silver  chloride, 
barium  sulphate  and  lead  sulphate  in  addition  to  fat.  Examine 
as  directed  under  "Metallic  Poisons  IV"  (see  page  163). 

H.  Thoms^  destroys  organic  matter  in  the  apparatus  shown 

in  Fig.  12.     Oxidation  is  carried  on  in  an  ordinary  fractioning 

flask  (A)  with  the  tubulus  (B)  bent  upward.     A  separating 

funnel  (C),  held  in  the  neck  of  the  flask  by  a  stopper,  contains 

an  aqueous  solution  of  potassium  chlorate  (i  :  20)  saturated  at 

room  temperature.     The  organic  matter  is  in  the  flask  as  a 

thin  mixture  with  12.5  per  cent,  hydrochloric  acid.     Add  about 

I  gram  of  solid  potassium  chlorate  and  warm  the  flask  on  a 

boiling  water-bath.     When  the  mass  in  the  flask  is  warm,  let 

the  potassium  chlorate  solution  run  in  drop  by  drop  and  shake 

constantly.     Care  must  be  taken  not  to  add  too  much  of  this 

solution  at  once;  otherwise  the  procedure  is  identical  with  that 

previously  described. 

Notes. — Potassium  chlorate  and  hydrochloric  acid  evolve  chlorine  (a  and  /3)) 
part  of  which  acts  upon  the  organic  material  and  part  in  contact  with  water  forms 
oxygen  and  oxygen  acids  of  chlorine  (HOCl)  (7  and  5)  which  are  strong  oxidizing 

(a)  KCIO3  +    HCl  =     HCIO3  +    KCl, 
(13)   HCIO3  +  SHCI  =  3CI2        +  3H2O, 
(7)   CI2       +    H2O  =  2HCI     +   o, 
(5)    CI2        +    HaO^    HOCl    +    HCl. 

White  Arsenic  (AS2O3)  in  a  mixture  probably  cannot  be  volatilized  as  arsenic 
trichloride  (AsClsj  in  the  procedure  described  but  is  oxidized  to  non-volatile 
arsenic  acid  (H3ASO4): 

AS2O3  +  2H2O  +  2CI2  =  AS2O5  +  4HCI, 
AS2O6  +  3H2O  =  2H3ASO4. 

There  always  remains,  even  after  the  most  thorough  treatment  with  hydro- 
chloric acid  and  potassium  chlorate,  an  insoluble  white  residue  whoUy  unafltected 
by  the  action  of  chlorine.  This  is  the  case,  especially  after  the  oxidation  of 
vegetable  substances  or  cadaveric  material.  This  treatment  converts  a  portion 
of  the  organic  matter  into  volatile  compounds  (chloranil?)  which  have  a  sharp 
odor  and  attack  the  mucous  membranes.  For  this  reason  destruction  of  organic 
matter  should  take  place  in  a  hood  with  a  good  draft. 

Treatment  as  described  with  hydrochloric  acid  and  potassium  chlorate  converts 
metallic  poisons  into  inorganic  salts,  usually  chlorides  and  sulphates.  These 
either  remain  in  solution  or  appear  as  precipitates  (AgCl  and  BaS04).  Protein 
substances,  present  in  all  animal  and  vegetable  organisms,  precipitate  many 

1  H.  Thoms,  "Einfiihrung  in  die  praktische Nahrungsmittel  Chemie," Leipzig, 
1899.     Published  by  S.  Hirzel,  Leipzig,  1899.     Abbildung  64,  page  153. 


144  DETECTION   OF   POISONS 

heavy  metals,  as  mercury,  silver,  lead,  copper  and  zinc  from  solutions  of  their 
salts.  These  metals  are  then  in  the  form  of  metallic  albuminates,  some  of  which 
dissolve  in  water  with  great  difficulty  and  are  very  stable.  Usually  these  metal- 
lic protein  compounds  must  receive  further  treatment  before  it  is  possible  to 
detect  the  metal.  Many  organic  acids,  as  tartaric  acid,  and  carbohydrates  inter- 
fere more  or  less  with  the  detection  of  heavy  metals.  In  combination  with  these 
organic  substances  heavy  metals  are  like  copper  in  potassium  cuprocyanide 
(K4Cu2(CN)6),  which  neither  sodium  hydroxide  nor  hydrogen  sulphide  will  pre- 
cipitate because  it  is  electrolytically  dissociated  in  solution  in  part  as  follows: 

K4Cu2(CN)6^4K-  +  Cu2(CN)6"". 

In  other  words,  the  solution  does  not  contain  cuprous  ions.  If  potassium  cupro- 
cyanide is  heated  with  hydrochloric  acid  and  potassium  chlorate,  copper  passes 
into  solution  as  cupric  chloride.  The  reagents  mentioned  above  now  precipitate 
copper,  for  cupric  chloride  ionizes  as  follows: 

CuCl2?^Cu'  -\-  2Cr. 

and  the  solution  now  contains  cupric  ions. 

The  detection,  therefore,  of  these  metallic  poisons  by  the  usual  ionic  reactions 
requires  a  prodecure  which  permits  the  analyst  to  bring  about  complete  destruc- 
tion of  interfering  organic  substances.  The  metals  in  question  are  thus  converted 
into  inorganic  salts. 

Potassium  chlorate  acts  best  only  in  strong  hydrochloric  acid  solution.  Conse- 
quently this  acid  should  always  be  in  excess.  If  the  mass  becomes  too  thick  at 
any  time  during  heating,  it  should  be  diluted  with  water  or  dilute  hydrochloric 
acid.  Also  the  contents  of  the  flask  should  be  well  shaken  during  treatment  with 
potassium  chlorate,  to  prevent  a  large  quantity  of  this  salt  from  collecting  upon 
the  bottom  of  the  flask.  Such  an  occurrence  may  cause  an  explosion  due  to 
formation  of  the  exceedingly  unstable  dioxide  of  chorine  (C102).^ 

The  author  employs  in  such  analyses  12.5  per  cent,  hydrochloric  acid  (sp.  gr. 
1. 061),  saturated  with  hydrogen  sulphide  and  kept  in  a  loosely  stoppered  bottle. 
This  insures  precipitation  of  the  final  traces  of  arsenic  sometimes  present  even  in 
the  purest  commercial  acid.  Before  being  used,  this  acid  is  filtered  through  ash- 
free  paper  to  remove  precipitated  sulphur  which  may  contain  arsenic  sulphide. 

Cadaveric  material,  heated  with  hydrochloric  acid  and  potassium  chlorate,  is 
dissolved  rather  easily.  An  experiment,  in  which  100  grams  of  stomach  and 
duodenum,  20  grams  of  stomach  contents,  75  grams  of  kidney  and  200  grams  of 
liver  (in  all  395  grams)  were  treated  as  described,  required  about  i  hour  for  com- 
plete solution.  The  insoluble  part  was  collected  upon  a  filter  and  washed.  It 
was  amorphous,  gummy,  yellowish  white  and  greasy.  After  being  dried  upon 
a  porous  earthen  plate,  it  weighed  52  grams.  Dried  at  100°,  it  weighed 
only  32  grams. 

2.  Sonnenschein-Jeserich  Method 

This  method  requires  the  use  of  pure  free  chloric  acid  instead 
of  potassium  chlorate.     Place  the  finely  divided  material  in  a 

1  (a)   KCIO3  -f  HCl  =  HCIO3  +  KCi, 
(/3)   3HCIO3  =  HCIO4  +  2CIO2  +  H2O. 


METALLIC   POISONS  145 

large  flask  and  dilute  with  water.  Add  a  few  cc.  of  chloric  acid 
and  warm  slowly  and  cautiously  ui)()n  the  water-bath.  As  soon 
as  the  mass  swells  and  becomes  porous,  gradually  add  small 
portions  of  hydrochloric  acid.  Even  a  considerable  quantity 
of  cadaveric  material  will  dissolve  in  2-3  hours.  Water  lost 
by  evaporation  should  be  replaced  occasionally,  otherwise  the 
reaction  may  take  place  with  explosive  violence.  In  other  re- 
spects, the  product  of  the  reaction  should  be  treated  as  already 
described. 

3.  C.  Mai's  Method  ^ 

Mix  the  finely  divided  material  with  dilute  hydrochloric  acid 
(i  :  12)  until  thin.  Add  a  little  potassium  chlorate  and  heat 
over  a  free  flame,  adding  from  time  to  time  small  quantities  of 
potassium  chlorate  (0.2  gram).  Cool  as  soon  as  liquefaction  of 
the  mass  is  complete.  Fat  separates  and  usually  can  be  re- 
moved easily  from  the  liquid.  Heat  this  fat  once  or  twice  with 
very  dilute  nitric  acid,  filter  and  add  the  filtrate  to  the  main  part 
of  the  liquid.  Continue  heating  the  latter,  adding  small  quan- 
tities of  ammonium  persulphate,  (H4N)2S208,,  until  the  liquid  is 
clear  and  light  yellow.  Filter  and  saturate  the  filtrate  as  usual 
with  hydrogen  sulphide.  Ammonium  persulphate  is  a  power- 
ful oxidizing  agent  and  also  adds  nothing  non-volatile  to  the 
liquid. 

Examination  of  Filtrate  for  Metallic  Poisons 
Precipitation  by  Hydrogen  Sulphide 

A  solution  properly  prepared  according  to  the  Fresenius-v. 
Babo  or  any  other  method,  freed  from  excess  of  hydrochloric 
acid  and  filtered,  should  have  only  a  faint  yellow  color.-  Heat 
such  a  solution  in  a  flask  upon  the  water-bath  and  saturate 
with  arsenic-free  hydrogen  sulphide.^     Pass  hydrogen  sulphide 

*  Zeitschrift  fiir  Untertersuchung  der  Nahrungs-  und  Genussmittel  5,  1106 
(1902). 

^Chromium  in  not  too  small  quantity  imparts  more  or  less  of  a  green 
color  both  to  the  solution  and  the  jfiltrate  from  the  hydrogen  sulphide  precipitate, 
owing  to  the  presence  of  chromic  chloride  (CrCls). 

^  Prepare  arsenic-free  hydrogen  sulphide  b}"  saturating  dilute  sodium  hydroxide 
solution  with  hydrogen  sulphide  from  crude  iron  sulphide  and  commercial  hydro- 
10 


146 


DETECTION   OF   POISONS 


for  0.5-1  hour  or  longer^  into  the  hot  solution  and  continue 
this  treatment  after  the  solution  has  been  removed  from  the 
water-bath  and  is  cold. 

Allow  the  solution  saturated  with  hydrogen  sulphide  to  stand 
in  the  loosely  stoppered  flask  for  several  hours  or  until  the  next 
day.  If  the  solution  then  smells  of  hydrogen  sulphide  and 
blackens  a  piece  of  lead  acetate  paper  held  over  it,  the  next  step 


Fig.  13. — Apparatus  for  Generating  Arsenic-free  Hydrogen  Sulphide,  (a) 
Generator  with  dilute  sulphuric  acid;  (5)  Separating  funnel  with  NaSH;  (c)  Wash- 
bottle;  (d)  Solution  to  be  saturated  with  H2S. 


in  the  process  may  be  taken.  Otherwise  warm  the  solution 
once  more  upon  the  water-bath  and  again  saturate  with  hydro- 
gen sulphide.  Finally  collect  the  hydrogen  sulphide  precipi- 
tate upon  a  small  paper  and  wash  with  hydrogen  sulphide  water. 
Examine  the  precipitate  for  arsenic,  antimony,  tin,  mercury, 
lead,  copper,  bismuth  and  cadmium  (Metallic  Poisons  I  and  II) 

chloric  acid.  Pour  this  sodium  hydro-sulphide  (NaSH)  solution  into  a  separating 
funnel  and  add  slowly  to  dilute  sulphuric  acid  (1:4).  The  generation  of  the  gas 
can  be  carried  on  in  the  apparatus  shown  in  Fig.  13. 

^  In  laboratory  experiments  treatment  with  hydrogen  sulphide  may  be  shortened 
somewhat.  A  Kipp  generator  in  which  the  gas  is  prepared  from  iron  sulphide 
and  hydrochloric  acid  may  be  used. 


METALLIC   POISONS  147 

and  the  filtrate  from  this  precipitate  for  chromium  and  zinc 
(MetaUic  Poisons  III). 

Vegetable  and  animal  substances,  after  treatment  with 
hydrochloric  acid  and  potassium  chlorate,  frequently  give 
liquids  yielding  colored  precipitates^  with  hydrogen  sulphide 
even  in  the  absence  of  the  metals  mentioned  above.  Such 
precipitates  consist  largely  of  organic  sulphur  compounds. 
Consequently,  if  hydrogen  sulphide  produces  such  a  colored 
precipitate  in  acid  solution,  it  is  not  final  proof  of  the  presence 
of  a  metallic  poison.  Also  without  further  examination  it  is 
impossible  to  decide  from  the  color  of  the  hydrogen  sulphide 
precipitate  as  to  the  presence  of  a  particular  metal. 

Complete  Precipitation.— Before  testing  for  chromium  and 
zinc  in  the  filtrate  from  the  hydrogen  sulphide  precipitate,  add 
about  lo  times  the  volume  of  strong  hydrogen  sulphide  water 
to  a  small  portion  of  the  solution,  stir  well  and  let  stand  several 
minutes.  Unless  a  colored  precipitate  appears,  the  metals  in 
question  (Metallic  Poisons  I  and  II)  have  been  completely 
removed  and  the  filtrate  may  then  be  further  tested  for  chrom- 
ium and  zinc  (Metallic  Poisons  III).  Otherwise,  first  dilute 
the  entire  filtrate  from  the  hydrogen  sulphide  precipitate  with 
water  and  again  saturate  with  hydrogen  sulphide.  In  presence 
of  much  hydrochloric  acid,  lead  and  cadmium  are  incompletely 
precipitated  by  hydrogen  sulphide  (see  above) . 

Treatment  of  Hydrogen  Sulphide  Precipitate  with  Ammonia 
and  Yellow  Ammonitmi  Sulphide. — Extract  the  thoroughly 
washed  hydrogen  sulphide  precipitate  while  still  moist  upon  the 
filter  with  a  hot  mixture  of  approximately  equal  parts  of  am- 
monia and  yellow  ammonium  sulphide.  Heat  about  5-10  cc. 
of  the  mixture  of  ammonia  and  yellow  ammonium  sulphide  to 
boiling  and  drop  the  solution  over  the  precipitate  upon  the  filter. 
Reheat  the  filtrate  and  again  pour  over  the  precipitate.  Repeat 
this  operation  several  times.     Finally  wash  the  filter  with   a 

^  Repeated  treatment  with  potassium  chlorate  and  hj-drochloric  acid  dissolves 
thoroughly  washed  casein  and  fibrin  almost  completel}'  and  gives  a  filtrate  from 
which  hydrogen  sulphide  precipitates  dirty  yellow  to  brownish  substances. 
These  products  are  amorphous  and  contain  organic  sulphur  compounds  together 
with  much  free  sulphur. 


148  DETECTION   OF   POISONS 

few  cc.  of  a  fresh  mixture  of  ammonia  and  yellow  ammonium 
sulphide.  Test  the  entire  filtrate  for  arsenic,  antimony,  tin 
and  copper^  =  Metallic  Poisons  I.  Test  the  residue  upon  the 
filter  for  mercury,  lead,  copper,  bismuth  and  cadmium  =  Me- 
tallic Poisons  II. 

METALLIC  POISONS  I 

Examination  of  the  Part  of  the  Hydrogen  Sulphide  Precipitate  Soluble 
in  Ammonia-Anmionimn  Sulphide 

Arsenic,  Antimony,  Tin,  Copper 

Use  the  solution  prepared  as  described  by  treating  the  hydro- 
gen sulphide  precipitate  with  a  hot  mixture  of  ammonia  and 
yellow  ammonium  sulphide.  This  solution  is  usually  dark 
brown  owing  to  dissolved  organic  substances.^  Evaporate  the 
solution  to  dryness  in  a  porcelain  dish  upon  the  water-bath. 
Moisten  the  cold  residue  with  fuming  nitric  acid  and  again 
evaporate.  Then  intimately  mix  the  residue  with  about  3  times 
its  volume^  of  a  mixture  of  2  parts  of  sodium  nitrate  and  i  part 
of  dry  sodium  carbonate.  Thoroughly  dry  this  mixture  upon 
the  water-bath  and  introduce  small  portions  at  a  time  into  a 
porcelain  crucible  containing  a  little  fused  sodium  nitrate 
heated  to  redness.     After  the  final  addition,  heat  the  crucible 

^  Copper  sulphide  (CuS)  is  somewhat  soluble  in  hot  yellow  ammonium  sul- 
phide. An  ammonium  sulphide  solution  containing  copper,  treated  as  described 
on  page  149,  yields  copper  oxide  which  gives  the  melt  a  more  or  less  gray  or 
black  appearance.  If  the  melt  is  extracted  with  water,  the  residue  contains 
black  copper  oxide  with  stannous  oxide  and  sodium  pyro-antimonate.  To  detect 
copper,  dissolve  the  black  residue  in  a  little  hot  dilute  hydrochloric  acid  and  divide 
the  solution  into  two  parts.  Add  ammonia  to  i  part  until  alkaline.  The  solu- 
tion is  blue,  if  copper  is  present.  Add  potassium  ferrocyanide  solution  to  the 
other  part.  A  brownish  red  precipitate  of  cupric  ferrocyanide  (Cu2Fe(CN)„) 
appears,  if  copper  is  present. 

^  In  absence  of  metals  the  appearance  of  dark  colored  precipitates  (see  above) , 
when  the  hydrochloric  acid  solution  is  treated  with  hydrogen  sulphide,  should  not 
be  misunderstood.  Such  precipitates  are  due  to  organic  substances  soluble  with 
a  dark  brown  color  in  a  hot  mixture  of  ammonia  and  yellow  ammonium  sulphide. 

^  In  most  laboratory  experiments  3  grams  of  a  mixture  of  2  grams  of  sodium 
nitrate  and  i  gram  of  sodium  carbonate  is  sufficient.  A  large  excess  of  sodium 
nitrate  should  be  avoided. 


METALLIC   POISr)NS  ]49 

a  short  time,  introducing  possibly  a  little  more  sodium  nitrate, 
until  tlie  fused  mass  is  colorless.  In  presence  of  copper  the 
melt  is  gray  or  grayish  black  from  copper  oxide.  Sodium 
arsenate,  sodium  pyro-antimonate,  sodium  stannate,  as  well  as 
stannic  oxide  and  copper  oxide,  may  also  be  present.  Soften 
the  cold  melt  with  hot  water  and  wash  into  a  flask.  Add  a 
little  acid  sodium  carbonate  to  the  clear  or  cloudy  liquid  to  de- 
compose the  small  quantity  of  sodium  stannate  possibly  in  solu- 
tion and  precipitate  all  the  tin  as  stannic  oxide  and  then  filter. 
The  filtrate  (A)  contains  any  arsenic  present  as  sodium  arsen- 
ate (Na2HAs04)  and  the  residue  upon  the  filter  (B)  may  con- 
tain^  sodium  pyro-antimonate  (NasHaSbaOr),  stannic  and  copper 
oxides. 

Examination  of  Filtrate  A  for  Arsenic 
Arsenic  is  isolated  as  the  element.  Positive  proof  of  the  pres- 
ence of  the  poison  is  thus  afforded.  Two  methods  are  in  use  for 
this  purpose,  namely,  the  Marsh-BerzeHus  and  the  Fresenius- 
V.  Babo  method.  Both  are  very  accurate  and  exclude  any 
confusion  of  arsenic  and  antimony. 

Chapter  V  gives  the  details  for  detecting  arsenic  by  the  very 
delicate  biological  test,  which  requires  the  use  of  certain  moulds, 
and  also  for  the  elctrolytic  separation  of  arsenic  at  the  cathode 
as  arsine. 

I.  Marsh-Berzelius  Method 

Principle. — Nascent  hydrogen  converts  oxygen  compounds 

of  arsenic,  arsenious  and  arsenic  acids,  as  well  as  arsenites  and 

arsenates  into  arsine,  AsHs  : 

AS2O3  -I-  12H  =  2ASH3  +  ^H.o, 

AS2O5.3H2O2  +  16H  =  2ASH3  +  8H2O. 

At  a  red  heat  arsine  is  decomposed  into  metallic  arsenic  and 
hydrogen: 

AsHs  =  As  -f-  3H. 

This  reaction  represents  the  formation  of  the  arsenic  mirror. 

1  Even  in  the  absence  of  the  substances  mentioned  under  B,  a  small  insoluble 
residue  usualty  appears.  This  may  come  from  the  porcelain  crucible,  the  glazing 
of  which  is  slightly  attacked  in  the  fusion  with  sodium  nitrate  and  carbonate. 

-AS2O5.3H2O  is  the  dualistic  method  of  writing  2H3ASO4. 


150  DETECTION   OF   POISONS 

Also  hydrogen  containing  arsine  burns  with  ,a  bluish  white 
flame  (a).  Depress  a  piece  of  cold  porcelain  upon  such  a  flame. 
Hydrogen  will  burn  but  a  deposition  of  metallic  arsenic  takes 
place  (j8).     This  is  the  so-called  arsenic  spot: 

(a)   sAsHs  +  3O2  =    AS2O3  +  3H2O, 

(;S)     2ASH3  +  30=    2AS  +  3H2O. 

Hydrogen  containing  arsine  precipitates  black  metalUc  silver, 
if  passed  into  dilute  silver  nitrate  solution.  The  solution  con- 
tains arsenious  acid: 

AsHs  +  3H2O  +  6AgN03  =  H3ASO3  +  6HNO3  +  6Ag. 

Procedure. — First  acidify  "  Filtrate  A,"  prepared  as  described 
(page  149)  and  possibly  containing  sodium  arsenate,  with  dilute 
arsenic-free  sulphuric  acid.  Evaporate  this  solution  in  a  porce- 
lain dish  upon  an  asbestos  plate  over  a  small  free  flame.  Add  a 
few  drops  of  concentrated  sulphuric  acid  to  expel  completely 
any  nitric  acid  possibly  preseiit  in  the  residue  and  heat  until 
copious  white  fumes  of  sulphuric  acid  appear.  The  residue^ 
in  the  porcelain  dish  is  a  thick  colorless  Hquid  having  a  strong 
acid  reaction.  Arsenic,  if  present,  is  in  the  form  of  arsenic  acid 
which  when  cold  frequently  solidifies  to  a  white  crystalline 
mass.  Examine  the  solution  of  this  residue  in  the  Marsh  ap- 
paratus for  arsenic.  The  same  solution  may  also  be  used  in 
testing  for  arsenic  electrolytically  (see  pages  226  and  231). 

Marsh  Apparatus. — Place  30-40  grams  of  pure  arsenic-free 
zinc^  (granulated  or  in  small  rods)  in  the  reduction  flask  A  of 
the  Marsh  apparatus  (Fig.  14).  Pour  cold  dilute  arsenic-free 
sulphuric  acid  upon  the  metal.  This  acid  should  contain 
15-16  per  cent,  of  H2S04.^  Control  the  temperature  of  the 
solution,  which  should  not  rise  much  during  the  analysis,  by 

^  To  insure  complete  removal  of  nitric  acid,  test  a  few  drops  of  this  residue  with 
ferrous  sulphate  and  sulphuric  acid. 

^The  passage  of  hydrogen  from  15-20  grams  of  zinc,  treated  with  dilute 
arsenic-free  sulphuric  acid,  through  the  strongly  heated  ignition  tube  C  of  the 
Marsh  apparatus  should  not  give  a  trace  of  arsenic  after  i  hour.  The  metal  is 
then  pure  enough  for  use.  Bertha  spelter  from  the  New  Jersey  Zinc  Company 
will  meet  such  a  test. 

^  Add  I  volume  of  pure  arsenic-free  concentrated  sulphuric  acid  to  5  volumes  of 
distilled  water.     This  diluted  acid  when  cold  is  suitable  for  use  in  the  M  arsh  test. 


METALLIC   POISONS 


151 


generating  hydrogen  slowly.  Otherwise,  there  is  danger  of 
partial  reduction  of  sulphuric  acid  to  sulphur  dioxide  and  then 
to  hydrogen  sulphide  which  interferes  more  or  less  with  the 
detection  of  arsenic.  Place  the  reduction  flask  A  in  a  dish  of 
cold  water,  if  the  acid  becomes  too  warm. 

Certain  precautions  are  necessary  in  using  the  Marsh  ap- 
paratus. 

1.  Have  the  apparatus  absolutely  tight. 

2.  Expel  air  completely  before  igniting  hydrogen.     To  tell 
when  this  point  is  reached,  collect  hydrogen  in  a  dry  test-tube 


Fig.  14. — Marsh  Apparatus,     (a)  Hydrogen-generator;  (6)  Chloride  of  cal- 
cium drying-tube;  (c)  Hard  glass  tube;  {d)  Arsenic  mirror. 


until  it  ignites  without  detonation  when  carried  to  a  flame.  If 
the  hydrogen  stands  this  test,  ignite  the  gas  at  the  end  of  igni- 
tion tube  C.  There  is  no  danger  of  an  explosion  within  the 
apparatus.  If  the  apparatus  is  tight  and  the  evolution  of 
hydrogen  is  not  too  rapid,  it  requires  about  8  minutes  to  expel 
the  air. 

3.  Test  the  hydrogen  to  insure  its  entire  freedom  from  arsenic. 
Neither  the  arsenic  mirror  nor  spot  appears. 

If  the  hydrogen  is  arsenic-free,  graduall}^  introduce  the 
perfectly  cold  sulphuric  acid  solution,  containing  arsenic  as 
arsenic  acid  (page  150),  in  small  portions  into  reduction  flask  A. 
At  the  same  time  heat  ignition  tube  C  to  redness  Just  back  of 


152  DETECTION   OF   POISONS 

the  capillary  tube.  If  the  solution  contains  arsenic,  the  gas 
generated  consists  of  a  mixture  of  arsine  (AsHs)  and  hydrogen. 
A  shining  mirror  of  metallic  arsenic  appears,  often  in  a  few 
minutes,  just  beyond  the  point  of  ignition.  Traces  of  arsenic 
require  considerable  time  before  a  brown  or  brownish  black 
film  appears.  A  piece  of  white  paper  held  behind  the  tube 
brings  out  clearly  even  a  minute  arsenic  mirror. 

Remove  the  flame  from  ignition  tube  C.  If  arsenic  is  present, 
the  hydrogen  flame  becomes  bluish  white.  At  the  same  time 
white  fumes  of  arsenious  oxide  (AS2O3)  arise  from  the  flame. 

To  produce  the  lustrous,  brownish  black  spot  (arsenic  spot), 
depress  a  cold  porcelain  dish  upon  the  hydrogen  flame. 

Arsine  has  an  exceedingly  characteristic,  garlic-like  odor. 
Extinguish  the  hydrogen  flame  and  allow  the  gas  to  escape.^ 
The  odor  is  evident  even  when  the  hydrogen  contains  traces 
of  arsine. 

A  third  method  of  detecting  arsenic  by  the  Marsh  apparatus 
consists  in  extinguishing  the  hydrogen  flame  and  passing  the 
gas  into  dilute  silver  nitrate  solution.  Arsine  darkens  this 
solution,  producing  a  black  precipitate  of  metallic  silver.  The 
solution  contains  arsenious  acid  and  free  nitric  acid.  (See 
reaction,  page  150.) 

Filter  through  a  double  paper  to  remove  silver  and  carefully 
neutralize  the  filtrate  with  a  few  drops  of  very  dilute  ammonium 
hydroxide  solution.  If  the  solution  is  neutral,  it  is  possible 
to  obtain  a  yellowish  white  precipitate  of  silver  arsenite 
(AgsAsOa)  but  this  compound  dissolves  easily  in  ammonium 
hydroxide  solution  and  in  nitric  acid. 

Extinguish  the  flame  at  the  end  of  reduction  tube  C  and  hold 
over  the  tube  a  strip  of  paper  moistened  with  concentrated 
silver  nitrate  solution  (1:1).  A  yellow  stain  appears,  if  the 
hydrogen  contains  arsine.  A  drop  of  water  added  to  this 
yellow  spot  changes  the  color  to  black.  This  is  Gutzeit's 
arsenic  test  (see  page  156). 

1  The  detection  of  arsine  by  other  tests  is  so  easy  that  it  seems  somewhat 
superfluous  to  confirm  its  presence  in  this  way  in  view  of  its  very  poisonous 
properties.     Tr. 


METALLIC   POISONS  153 

Differences  Between  Arsenic  and  Antimony  Spots  and  Mirrors 

Nascent  hydrogen  reduces  various  antimony  compounds  (SbClj,  SbjO?, 
HSbOs,  KSbOC4H40o,  etc.)  producing  the  colorless  gas  stibine  (Sblfs).  The 
behavior  of  this  compound  in  the  Marsh  api)aratus  closely  resembles  that  of 
arsine,  for  it  gives  a  spot  and  mirror,  and  precipitates  black  silver  antimonide 
(AgsSb)  but  not  metallic  silver,  if  passed  into  silver  nitrate  solution. 

The  procedure  employed  in  preparing  material  for  the  A^arsh  test  (see  page 
148)  separates  arsenic  from  antimony  and  excludes  the  possibility  of  the  two  met- 
als appearing  in  the  Marsh  test  at  the  same  time.  Since  the  identification  of 
arsenic  mirrors  and  spots  by  othef  tests  is  important,  the  differences  between  ar- 
senic and  antimony  should  be  pointed  out.  The  suspicion  that  antimony  is 
present  often  necessitates  other  confirmatory  tests  (see  Antimony,  page  157). 
Introduce  the  solution  into  the  Marsh  apparatus  and  produce  the  antimony 
spot  and  mirror. 

The  differences  between  arsenic  and  antimony  spots  and  mirrors  are: 

1.  The  arsenic  mirror  has  a  high  metallic  luster.  It  is  brownish  black  and 
volatile.  Owing  to  this  latter  property,  it  sublimes  easily  when  heated  in  a 
stream  of  hydrogen.  In  the  case  of  the  antimony  mirror,  which  appears  on  both 
sides  of  the  flame,  the  metal  in  contact  with  the  heated  glass  fuses  and  is  silver 
white.  But  in  those  places  removed  from  the  flame  it  is  almost  black  and  has 
hardly  any  luster.  Stibine  decomposes  at  a  temperature  much  below  that 
required  for  arsine.  This  fact  explains  the  deposition  of  this  metal  on  both  sides 
of  the  flame.  Antimony  volatilizes  at  a  high  temperature  and  consequently 
sublimes  with  difficulty. 

2.  The  arsenic  spot,  if  not  too  heavy,  is  brownish  black  or  brown  and  lustrous. 
It  dissolves  readily  in  sodium  hypochlorite  solution,  forming  arsenious  acid: 

3H2O  +  2As  +  3NaOCl  =  2H3ASO3  +  sNaCl. 

The  antimony  spot  is  dull,  velvet  black  and  without  luster.  A  thin  film  of 
antimony  is  never  brown  but  has  a  dark,  graphite-like  appearance.  It  is  insolu- 
ble in  sodium  hypochlorite  solution. 

3.  A  drop  of  concentrated  nitric  acid,  or  moist  chlorine,  at  once  dissolves  the 
arsenic  spot  forming  arsenic  acid.  Neutralize  with  ammonia  and  add  silver 
nitrate  solution.     A  reddish  precipitate  of  silver  arsenate  (Ag3As04)  appears. 

Nitric  acid,  or  moist  chlorine,  also  dissolves  the  antimony  spot  but  silver 
nitrate  does  not  produce  a  colored  precipitate. 

4.  Gently  heat  the  ignition  tube  and  pass  a  stream  of  dry  hj-drogen  sulphide 
over  the  arsenic  mirror.  Yellow  arsenic  trisulphide  (AS2S3)  appears.  The 
antimony  mirror  becomes  brownish  red  to  black  (,Sb2S3). 

5.  Arsine  passed  into  silver  nitrate  solution  precipitates  black  metallic  silver 
and  the  filtrate  from  such  a  precipitate  contains  arsenious  acid.  But  stibine 
precipitates  black  silver  antimonide  (Ag3Sb)  and  the  filtrate  does  not  contain  a 
trace  of  antimony  since  the  precipitation  of  black  AgsSb  is  complete.  To  detect 
antimony,  collect  the  black  precipitate  upon  paper,  wash  and  heat  for  some  time 
in  10-15  P^r  cent,  tartaric  acid  solution.  Antimony  dissolves,  whereas  silver 
remains  as  a  grayish  white  residue.  Add  dilute  hj-drochloric  acid  to  this  solu- 
tion and  then  treat  with  hydrogen  sulphide.  Antimony  appears  as  orange-red 
antimony  trisulphide. 


154 


DETECTION   OF  POISONS 


2.  Fresenius-von  Babo  Method 

Principle. — Fusion  of  oxygen  and  sulphur  compounds  of 
arsenic  with  a  mixture  of  sodium  carbonate  and  potassium 
cyanide  causes  reduction  with  formation  of  an  arsenic  mirror. 
As  a  result  potassium  cyanide  changes  to  potassium  cyanate 
(KCNO)  or  potassium  sulphocyanate  (KSCN) : 

AS2O3  +  3KCN  =  As2  +  3KCNO, 

AS2S3  +  3KCN  =  As2  +  3KSCN. 

Procedure. — Use  for  this  test  the  sulphuric  acid  solution 
prepared  by  the  method  already  described  and  containing  ar- 
senic in  the  form  of  arsenic  acid  (page  150).  To  reduce  arsenic 
acid  to  arsenious  acid,  add  a  few  cc.  of  sulphurous  acid  to  the 
solution  and  heat  until  the  odor  of  this  acid  has  disappeared. 


Fig.  15. — Fresenius-Von  Babo  Apparatus,  (a)  Carbon  dioxide  generator; 
(6)  Drying-bottle  with  pure,  concentrated  sulphuric  acid;  (c)  Ignition-tube  and 
boat. 

Dilute  this  solution  with  water  and  treat  with  hydrogen  sul- 
phide. Collect  the  precipitate  of  arsenic  trisulphide  (AS2S3) 
upon  a  small  filter  and  wash  thoroughly.  Dissolve  the  pre- 
cipitate upon  the  filter  in  a  little  hot  ammonium  hydroxide  solu- 
tion. Evaporate  this  solution  in  a  porcelain  dish  upon  the 
water-bath  and  heat  the  residue  with  concentrated  nitric  acid. 
Expel  the  latter  completely  by  evaporation,  moisten  the  residue 


METALLIC   POISONS  155 

with  a  little  water  and  add  enough  dry  sodium  carbonate  to 
render  the  mixture  distinctly  alkaline.  Dry  thoroughly  upon 
the  water-bath  and  triturate  the  residue  in  a  mortar  with  several 
times  the  quantity  of  a  mixture  of  3  parts  of  dry  sodium  car- 
bonate and  I  part  of  pure  potassium  cyanide. 

Transfer  this  mixture  to  a  porcelain  boat  and  place  in  an  igni- 
tion tube  of  hard  glass.  Heat  in  a  stream  of  carbon  dioxide 
(Fig.  15)  dried  by  means  of  arsenic-free  sulphuric  acid. 

To  expel  moisture,  first  heat  the  ignition  tube  gently  where 
the  boat  is  and  then  ignite  at  a  bright  red  heat.  A  mirror 
of  arsenic  appears  upon  the  cooler  part  of  the  tube,  if  arsenic  is 
present. 

A  simpler  method  of  detecting  arsenic  by  means  of  potassium 
cyanide  is  often  used.     Heat  the  thoroughly  dried  material 


A. 

Fig.  16. — (A)  Substance  and  fusion-mixture;  (B)  Arsenic  mirror. 

containing  arsenic  (AS2O3,  AS2S3)  in  a  bulb  tube  with  a  dry  mix- 
ture of  sodium  carbonate  and  potassium  cyanide  until  fusion 
takes  place.  If  the  tube  is  smaller  above  the  bulb,  the  arsenic 
mirror  will  form  in  the  constricted  area  (Fig.  16). 

Other  Arsenic  Tests 

I.  Bettendorff's  Test. — Concentrated  stannous  chloride  solu- 
tion precipitates  metallic  arsenic  from  arsenious  acid  cold  and 
from  arsenic  acid  with  heat  or  after  long  standing.  This  test 
requires  the  use  of  a  special  stannous  chloride  solution.^  The 
solution  is  red  to  brownish  red,  if  only  traces  of  arsenic  are  pres- 
ent. .  More  than  traces  of  arsenic  produce  a  black  precipitate  of 
arsenic : 

AS2O3  +    6HC1  -i-  sSnClo  =  2As  -|-  3H2O  +  sSnCU, 

AssOb  -I-  loHCl  +  sSnCla  +  3H2O  =  2As  +  8H2O  +  sSnCh. 

Use  for  this  test  the  sulphuric  acid  solution  obtained  as  de- 
scribed above  (page  150)  which  contains  arsenic  in  the  form  of 

^  See  page  315  for  the  preparation  of  this  reagent. 


156  DETECTION   OF   POISONS 

arsenic  add.  Bettendorff  s  test  is  not  as  delicate  as  the  Marsh 
test. 

2.  Gutzeit's  Test.^ — This  test  permits  the  detection  of  minute 
traces  of  arsenious  and  arsenic  acids,  as  well  as  their  salts,  with 
certainty.  Generate  hydrogen  in  a  test-tube  from  arsenic-free 
zinc  and  pure  dilute  hydrochloric  acid.  To  remove  any  sul- 
phurous acid  or  hydrogen  sulphide,  add  a  few  drops  of  iodine 
solution  until  the  liquid  is  yellow.  Then  add  the  solution  to  be 
tested  and  place  a  loose  cotton  plug  in  the  neck  of  the  test-tube. 
If  arsenic  is  present,  the  silver  nitrate  spot  becomes  lemon- 
yellow. 

AsHs  +  6AgN03  =  (sAgNOs.AgaAs)  +  3HNO3. 

Gradually  a  brownish  black  border  forms  around  the  yellow 
spot.  A  drop  of  water  at  once  turns  the  spot  black  from  sepa- 
ration of  metallic  silver. 

(sAgNOs.AgsAs)  +  3H2O  =  6Ag  +  H3ASO3  +  3HNO3. 

This  is  a  very  delicate  arsenic  test.  One  drop  of  o.i  per  cent, 
potassium  arsenite  solution  produces  a  distinct  yellow  color 
upon  the  silver  paper.  Gutzeit's  test  is  positive  with  even  0.05 
milligram  of  As203.^  The  sulphuric  acid  solution  containing 
arsenic  as  arsenic  acid  (see  page  150)  may  be  used  for  this  test. 

But  Gutzeit's  test  is  not  as  characteristic  of  arsenic  as  the 
Marsh  test.  Stibine,  phosphine  from  phosphorus  in  zinc  and 
even  hydrocarbons  color  the  silver  nitrate  paper.  Dry  hydro- 
gen sulphide  also  produces  a  yellow  or  yellowish  green  spot  upon 
paper  moistened  with  concentrated  silver  nitrate  solution. 
The  latter  spot  has  a  black  border  which  gradually  extends 
until  the  entire  spot  becomes  black.  Poleck  gives  this  yellow 
compound  the  composition  (AgN03.Ag2S). 

Detection  of  Antimony,  Tin  and  Copper  in  Residue  B 

Residue  B  (see  page  149),  insoluble  in  water  and  obtained 
from  the  fusion,  may  contain  sodium  pyro-antimonate,  stannic 

^  Gutzeit,  Pharmazeutische  Zeitung  1879,  263;  and  Poleck  and  Ttimmel, 
Berichte  der  Deutschen  chemischen  Gesellschaft  16,  2435  (1883). 

2  See  page  233  for  the  application  of  this  method  to  the  quantitative  estimation 
of  arsenic.     Tr. 


METALLIC   POISONS  157 

and  cupric  oxides.  Treat  this  residue  upon  the  filter  with  a 
little  hot  dilute  hydrochloric  acid  (equal  parts  of  concentrated 
acid  and  water).  Pass  this  acid  repeatedly  through  the  paper 
until  most  of  the  residue  is  dissolved.  If  the  original  color 
of  residue  B  and  of  the  melt  was  gray  or  black,  first  examine  a 
portion  of  the  hydrochloric  acid  solution  for  copper.  Excess 
of  ammonia  produces  a  blue  color.  Potassium  ferrocyanide 
solution  gives  a  brownish  red  precipitate,  or  only  a  coloration 
with  traces  of  copper. 

Concentrate  the  remainder  of  the  hydrochloric  acid  solution 
to  a  few  drops  in  a  porcelain  dish  upon  the  water-bath,  and  put 
2  drops  of  this  solution  upon  platinum  foil  in  contact  with  zinc. 
Antimony  produces  a  black,  tin  a  grayish  and  copper  a  dark 
reddish  brown  spot  upon  platinum.  There  is  little  chance  of 
confusing  the  tin  or  copper  spot  with  that  given  by  antimony. 

Dilute  the  remainder  of  the  hydrochloric  acid  solution  with 
water  and  introduce  a  piece  of  zinc.  Keep  the  zinc  in  the  solu- 
tion, as  long  as  hydrogen  is  evolved.  Collect  the  black,  metal- 
lic flocks  from  this  operation  upon  a  small  filter.  Wash  thor- 
oughly, and  gently  warm  with  a  little  concentrated  hydrochloric 
acid.  Finally,  filter  the  solution.  Antimony  does  not  dissolve, 
whereas  tin  passes  into  solution  as  stannous  chloride  (SnClo), 
and  is  in  the  filtrate.  Apply  the  tests  described  under  tin  to 
this  solution. 


TIN 


The  solution,  treated  as  described  in  the  above  analytical 
procedure,  contains  tin  as  stannous  chloride  (SnCl-:).  Apply 
the  following  tests  for  this  metal: 

{a)  Merciiry  Test. — Add  to  a  portion  of  the  filtrate  a  few 
drops  of  mercuric  chloride  solution.  Tin  precipitates  white 
mercurous  chloride  (calomel).  Heat  produces  in  addition 
gray,  metallic  mercury,  if  there  is  a  large  excess  of  stannous 
chloride. 

(b)  Prussian  Blue  Test. — Add  to  a  second  portion  of  the 
filtrate  a  few  drops  of  a  dilute  mixture  of  ferric  chloride  and 


158  DETECTION   OF   POISONS 

potassium  ferricyanide  solutions.     Tin  produces  a  precipitate 
of  Prussian  blue. 

This  test  is  not  characteristic  of  tin,  as  many  other  substances  capable  of 
reducing  ferri-ferricyanide  to  ferri-ferrocyanide,  that  is  to  say,  to  Prussian  blue 
act  in  the  same  way. 

To  identify  antimony  further,  dissolve  the  black  flocks,  in- 
soluble in  hydrochloric  acid,  in  a  few  drops  of  hot  aqua  regia. 
Expel  excess  of  acid  upon  the  water-bath  and  dilute  the  residue 
with  water.  If  the  quantity  of  antimony  is  not  too  small,  water 
precipitates  white  antimony  oxychloride  (SbOCl).  Redissolve 
this  precipitate  in  a  little  dilute  hydrochloric  acid.  Test  a 
portion  of  this  solution  for  antimony  with  hydrogen  sulphide. 
Introduce  the  remainder  into  the  Marsh  apparatus  and  produce 
the  antimony  spot  and  mirror,  or  test  for  stibine  with  silver 
nitrate  solution  as  described  (see  page  153). 

METALLIC  POISONS  II 

Detection  of  Metals  Whose  Sulphides  are  Insoluble  in  Ammonitim  Sul- 
phide 
Mercury  Bismuth 

Lead  Copper 

Cadmiiim 

That  portion  of  the  hydrogen  sulphide  precipitate,  insoluble 
in  ammonium  sulphide  solution,  may  contain  mercury,  lead, 
bismuth,  copper  and  cadmium  sulphides.  Examine  this  pre- 
cipitate according  to  the  methods  employed  in  qualitative 
analysis.  Treat  a  small  precipitate  repeatedly  upon  the  filter 
with  a  few  cc.  of  warm,  rather  dilute  nitric  acid  (i  volume  of 
concentrated  acid  and  2  volumes  of  water).  Mercuric  sulphide 
does  not  dissolve,  but  the  other  sulphides  pass  into  solution  as 
nitrates. 

Detection  of  Merctiry  in  the  Residue  Insoluble  in  Nitric  Acid 

Always  examine  that  portion  of  the  hydrogen  sulphide  pre- 
cipitate, insoluble  in  nitric  acid,  for  mercury,  even  when  not 
black!     Treat  this  residue  upon  the  filter  with  a  little  hot, 


METALLIC    POLSONS  159 

somewhat  diluted  hydrochloric  acid,  containing  in  solution  a 
few  crystals  of  potassium  chlorate  and  pass  the  acid  through  the 
paper  several  times.  Evaporate  the  filtrate  to  dryness  in  a 
porcelain  dish  upon  the  water-bath,  and  dissolve  the  residue  in 
2-3  cc.  of  water  containing  hydrochloric  acid.  Filter  this  solu- 
tion, and  examine  the  filtrate  for  mercury. 

(a)  Stannous  Chloride  Test. — Add  to  a  portion  of  the  filtrate 
a  few  drops  of  stannous  chloride  solution.  A  white  precipitate 
of  mercurous  chloride  (calomel)  appears,  if  mercury  is  present. 
Excess  of  stannous  chloride,  especially  if  heat  is  applied,  re- 
duces this  precipitate  to  gray,  metallic  mercury. 

(b)  Copper  Test. — Put  a  few  drops  of  the  filtrate  upon  a 
small  piece  of  bright  copper.  Mercury  immediately  deposits 
a  gray  spot  which  has  a  silvery  luster  when  rubbed.  Wash 
the  copper,  upon  which  mercury  has  been  deposited,  successively 
in  water,  alcohol  and  ether.  Dry  thoroughly  and  heat  in  a 
small  bulb-tube  of  hard  glass.  Mercury  sublimes  and  collects 
in  small,  metallic  globules  on  the  cool  sides  of  the  tube.  A 
trace  of  iodine  vapor,  introduced  into  the  tube  immediately  trans- 
forms the  gray  sublimate  into  scarlet  mercuric  iodide  (Hgl2) . 

(c)  Phosphorous  Acid  Test. — Add  to  another  portion  of  the 
filtrate  some  phosphorous  acid  and  warm  gently,  A  white 
precipitate  of  mercurous  chloride  (calomel)  appears,  if  mercury 
is  present: 

2HgCl2  +  H2O  +  H3PO3    =   Hg2Cl2  +   2HCI  +  H3PO4. 

(d)  Precipitation  of  Mercuric  lodide.^ — Add  1-2  drops  of  very 
dilute  potassium  iodide  solution  to  the  remainder  of  the  filtrate. 
A  red  precipitate  (Hgl2),  readily  soluble  in  excess  of  potassium 
iodide,  shows  mercury: 

1.  HgCl2  +  2KI  =  Hglo  +  2KCI, 

2.  Hgl2  +  2KI  =  K2HgT4. 

Examination  of  the  Nitric  Acid  Solution 

The  nitric  acid  solution  may  contain  lead,  bismuth,  copper 
and  cadmium  nitrates.  Evaporate  this  solution  in  a  porcelain 
dish  nearly  to  dryness  and  dissolve  the  residue  in  a  Httle  hot 


160  DETECTION   OF   POISONS 

water.  If  the  solution  contains  lead,  dilute  sulphuric  acid  pro- 
duces a  heavy  white  precipitate  of  lead  sulphate.  Test  the 
filtrate  from  this  precipitate  for  bismuth,  copper  and  cadmium. 

(a)  Copper  and  Bismuth  Tests. — Excess  of  ammonium  hy- 
droxide solution,  added  to  most  of  this  filtrate,  produces  a  blue 
color  if  copper  is  present.  A  white  precipitate  at  the  same  time 
may  be  bismuthous  hydroxide^  (Bi(0H)3).  To  detect  bismuth, 
wash  the  precipitate  and  dissolve  upon  the  filter  in  a  few  drops 
of  hot  dilute  hydrochloric  acid.  Pour  this  solution  into  consid- 
erable water.  A  white  precipitate  of  bismuthous  oxychloride 
(BiOCl)  proves  the  presence  of  bismuth.  As  an  alternative 
test,  add  stannous  chloride  to  the  hydrochloric  acid  solution  and 
then  excess  of  sodium  hydroxide  solution.  A  black  precipitate 
of  metallic  bismuth  appears. 

(b)  Potassium  Ferrocyanide  Test,^ — Potassium  ferrocyanide 
solution  precipitates  copper  as  brownish  red  cupric  ferrocyanide 
(Cu2Fe(CN)  e)  •  Traces  of  copper  produce  a  brownish  red  color. 
There  is  a  deposit  of  cupric  ferrocyanide  after  some  time. 

(c)  Precipitation  of  Metallic  Copper. — A  bright  knife  blade, 
or  a  bright  iron  nail,  immersed  for  a  short  time  in  a  copper  solu- 
tion, becomes  red  from  a  coating  of  metallic  copper. 

To  detect  cadmium  in  presence  of  copper,  add  solid  potas- 
sium cyanide  to  the  blue  solution,  produced  by  ammonium 
hydroxide,  until  the  blue  color  is  discharged.  Then  pass  hydro- 
gen sulphide  through  the  solution.  Cadmium  is  precipitated  as 
the  yellow  sulphide  (CdS),  whereas  copper  remains  in  solution 
as  K4Cu2(CN)6,  potassium  cuprocyanide. 
.  When  copper  is  absent,  test  for  cadmium  by  passing  hydrogen 
sulphide  at  once  into  the  ammoniacal  solution.  If  a  reddish 
or  brownish  instead  of  a  yellow  precipitate  appears,  filter,  dry 
the  precipitate  upon  the  paper  and  heat  upon  charcoal  in  the 
blow-pipe  flame.     Cadmium  gives  a  brown  coating. 

^  Ammonium  hydroxide  solution  does  not  precipitate  pure  bismuthous  hydrox- 
ide (Bi(0H)3)  from  solutions  of  bismuth  salts  but  a  basic  salt,  the  composition 
of  which  depends  upon  the  temperature  and  concentration  of  the  particular 
solutions. 


METALLIC    POISONS  161 

METALLIC  POISONS  III 

Detection  of  Chromium  and  Zinc 

Test  the  filtrate  from  the  hydrogen  sulphide  precipitate  for 
chromium  and  zinc.  Concentrate  the  filtrate  to  about  one- 
third  its  original  volume  and  divide  this  solution  into  two  parts. 

Detection  of  Zinc 

Add  enough  ammonium  hydroxide  solution  to  render  one- 
half  the  concentrated  filtrate  alkaline.  This  treatment  usually 
gives  the  solution  a  dark  color.  Then  add  ammonium  sulphide 
solution  in  excess.  This  reagent  almost  always  produces  a 
precipitate  even  when  zinc  is  not  present,  since  solutions,  pre- 
pared from  animal  and  vegetable  materials,  usually  contain 
iron  compounds  and  phosphates  of  the  metals  of  the  earths  and 
of  the  alkaline  earths.  When  this  precipitate  has  settled,  add 
acetic  acid  until  the  solution  has  a  faint  acid  reaction.  Stir  the 
mixture  thoroughly,  and  allow  it  to  stand  for  some  time.  The 
color  of  the  precipitate  becomes  lighter,  because  acetic  acid  dis- 
solves sulphide  of  iron.  Moreover,  the  phosphates  are  partly 
dissolved,  except  ferric  phosphate  (FeP04)  which  is  insoluble  in 
acetic  acid.  Collect  the  precipitate  upon  a  filter.  Wash,  dry 
and  ignite  precipitate  and  filter  in  a  porcelain  crucible.  Before 
ignition,  moisten  the  filter  with  concentrated  ammonium  ni- 
trate solution.  Extract  the  residue  from  ignition  with  3  cc. 
of  boiling,  dilute  sulphuric  acid.  Filter,  and  divide  the  filtrate 
into  two  parts. 

(a)  Add  sodium  hydroxide  solution  in  excess  to  one  portion 
of  the  filtrate  and  shake  thoroughly.  Filter,  to  remove  the 
white  precipitate  of  ferric  phosphate  which  usually  appears,  and 
add  a  few  drops  of  ammonium  or  hydrogen  sulphide  solution  to 
the  clear  filtrate  and  heat.  Zinc,  if  present,  gives  a  white, 
flocculent  precipitate  of  zinc  sulphide. 

(b)  Add  ammonium  hydroxide  solution  in  considerable  ex- 
cess to  the  second  part  of  the  filtrate.     Filter,  to  remove  ferric 

phosphate,  and  acidify  the  filtrate  with  acetic   acid.     Warm 
11 


162  DETECTION   OF   POISONS 

the  solution  and  treat  with  hydrogen  sulphide.     Zinc,  if  present, 
appears  as  the  white  sulphide. 

(c)  Test  further  for  zinc  by  dissolving  the  precipitate  pro- 
duced by  ammonium  or  hydrogen  sulphide  (as  described  above 
in  a  and  h),  after  it  has  been  collected  upon  a  filter  and  thor- 
oughly washed,  in  a  few  drops  of  hot  dilute  hydrochloric  acid. 
Boil  until  hydrogen  sulphide  is  expelled,  and  filter  to  remove 
precipitated  sulphur.  Add  potassium  ferrocyanide  solution  to 
the  clear,  cold  filtrate.  This  precipitates  Zn2Fe(CN)6,  zinc 
ferrocyanide,  which  is  white,  slimy  and  nearly  insoluble  in  dilute 
hydrochloric  acid.^ 

Detection  of  Chromium 

To  test  for  chromium^  in  the  second  part  of  the  filtrate 
from  the  hydrogen  sulphide  precipitate,  concentrate  the  solu- 
tion in  a  porcelain  dish  to  a  small  volume.  Then  add  twice  the 
quantity  of  potassium  nitrate  and  sodium  carbonate,  until  the 
reaction  is  decidedly  alkaline.  Finally,  heat  this  mixture  until 
perfectly  dry.  Add  this  dry  residue  in  small  portions  to  a  little 
potassium  nitrate  fused  in  a  crucible.  In  fusing  a  large  quan- 
tity of  material,  it  is  advisable  to  use  a  large,  bright,  nickel 
crucible  which  is  especially  adapted  for  this  operation.  When 
fusion  is  complete,  cool  thoroughly,  boil  crucible  and  contents 
with  water  in  a  porcelain  dish  and  filter  the  solution.  Chro- 
mium colors  the  filtrate  more  or  less  yellow.  Even  mere  traces' 
of  chromium  color  the  filtrate  yellow.  When  the  solution  of 
the  melt  is  colorless,  it  is  unnecessary  to  test  for  chromium.  To 
detect  chromium  when  the  filtrate  is  yellow,  divide  the  solution 
into  two  portions  and  make  the  following  tests : 

1  Excess  of  potassium  ferrocyanide  combines  with  zinc  ferrocyanide,  which  is 
first  precipitated,  and  forms  insoluble  potassium  zinc  ferrocyanide: 

3Zn2Fe(CN)6  +  K4Fe(CN)6  =  2K2Zn3(Fe(CN)6)2. 

2  In  testing  for  metallic  poisons,  chromic  oxide  (Cr203),  which  is  insoluble  in 
acids,  may  be  disregarded  as  it  is  not  poisonous. 

^  Two  drops  of  lo  per  cent,  potassium  chromate  solution  ( =  o  .oi  gram  of 
K2Cr04)  in  500  cc.  of  water  produce  a  marked  yellow  color.  Fifty  cc.  of  this 
solution  contain  o  .001  gram  of  K2Cr04  which  can  still  be  recognized  by  the 
yellow  color. 


mktallk:  poisons  103 

{a)  Chrome  Yellow  Test. — Add  acetic  acid  in  excess  to 
one  portion  of  the  filtrate,  and  boil  for  some  time  to  expel 
carbon  dioxide  and  nitrous  acid  as  completely  as  possible. 
Then  add  a  few  drops  of  lead  acetate  solution.  A  yellow 
precipitate  of  lead  chromate  (PbCrO^,  "Chrome  Yellow") 
appears,  if  chromium  is  present.  When  the  precipitate  is 
mixed  with  considerable  lead  sulphate  or  chloride,  the  color 
is  only  yellowish.  A  white  precipitate  is  due  to  PbS04,  PbCl2 
or  Pb3(P04)2.  When  the  aqueous  solution  of  the  melt  is  color- 
less, such  a  precipitate  is  usually  obtained. 

Potassium  nitrate,  always  present  in  the  melt,  lead  acetate  and  acetic  acid, 
brought  together  in  solution,  produce  a  distinct  yellow  color,  in  which  a 
white  precipitate  may  appear  yellow.  To  eliminate  this  source  of  error, 
allow  the  precipitate  to  settle,  collect  upon  a  filter  and  wash  thoroughly.  If 
it  is  pure  white,  chromium  is  absent. 

{h)  Reduction  Test. — Add  sulphurous  acid  to  the  second 
portion  of  the  yellow  filtrate.  The  yellow  color  changes  to 
green,  or  greenish  blue,  with  formation  of  chrome  alum.  This  is 
not  as  delicate  as  the  preceding  test. 

METALLIC  POISONS  IV 

Detection  of  Barium,  Lead  and  Silver  in  the  Residue  from  Hydrochloric 
Acid  and  Potassium  Chlorate 

Wash  the  residue  left  undissolved  in  the  treatment  with 
hydrochloric  acid  and  potassium  chlorate  thoroughly  with  water 
and  dry  in  the  air  closet  or  upon  a  porous  plate.  Then  add 
three  times  the  quantity  of  a  mixture  of  2  parts  of  potassium 
nitrate  and  i  part  of  sodium  carbonate  and  triturate  in  a  mortar 
with  the  filter.  Gradually  introduce  this  mixture  into  a  hot 
porcelain  crucible.  In  this  operation  organic  substances  (fats, 
fatty  acids,  etc.)  are  oxidized  by  potassium  nitrate  with  con- 
siderable deflagration.  Finally  when  all  the  material  is  in  the 
crucible,  add  0.25-0.5  gram  more  of  potassium  nitrate.  Cool 
the  melt,  soften  with  water,  wash  into  a  flask  and  pass  carbon 
dioxide  through  the  turbid  liquid  for  several  minutes.  This 
treatment  converts  caustic  alkali  into  carbonate  and  completely 
precipitates  lead  which  may  be  in  solution.     Then  heat  the 


164  DETECTION    OF    POISONS 

solution  to  boiling  and  let  settle  for  some  time.  Collect  upon 
paper  the  sediment^  which  may  contain  barium  carbonate,  basic 
lead  carbonate  and  metallic  silver.  Silver  gives  the  sediment 
a  gray  color.  Thoroughly  wash  the  precipitate  with  hot  water 
and  dissolve  upon  the  paper  in  hot,  rather  dilute  nitric  acid,^ 
passing  the  acid  through  the  paper  several  times.  Evaporate 
this  solution  to  dryness.  Dissolve  the  residue  in  water,  heat 
the  entire  solution  to  boiling,  and  precipitate  silver  with  dilute 
hydrochloric  acid.  Filter,  to  remove  silver  chloride,  and  pass 
hydrogen  sulphide  through  the  filtrate  to  precipitate  lead. 
To  test  for  barium  in  the  filtrate  from  lead  sulphide,  first  boil 
to  expel  hydrogen  sulphide  and  filter  to  remove  insoluble 
matter.  Then  add  dilute  sulphuric  acid  which  precipitates 
barium  sulphate. 

To  identify  silver  further,  dry  the  hydrochloric  acid  precipi- 
tate and  fuse  in  a  porcelain  crucible  with  a  little  potassium  cya- 
nide. Extract  the  melt  with  hot  water.  Metallic  silver  re- 
mains undissolved. 

To  confirm  the  presence  of  lead,  dissolve  the  hydrogen  sul- 
phide precipitate  in  hot  nitric  acid  and  evaporate  the  solution 
to  dryness.  Dissolve  the  residue  in  water,  filter  and  test  the 
solution  for  lead  with  sulphuric  acid  or  potassium  chromate. 

To  identify  barium  further,  collect  the  sulphuric  acid  pre- 
cipitate upon  paper,  thoroughly  wash  and  test  upon  a  clean 
platinum  wire  in  a  non-luminous  flame.  Barium  imparts  a 
yellowish  green  color  to  the  flame.  To  avoid  any  mistake, 
examine  this  flame  with  the  spectroscope.  These  reactions 
of  identification  are  always  necessary  in  toxicological  analysis. 

SYNOPSIS  OF  GROUP  HI 

Heat  the  residue  from  distillation  of  volatile  poisons,  or  a 
portion  of  original  material,  in  a  glass  flask  or  porcelain  dish 

1  Even  in  the  absence  of  barium,  lead  and  silver,  such  a  sediment  nearly 
always  appears.  In  that  case  it  usually  consists  of  the  material  of  the  porcelain 
crucible  the  glazing  of  which  is  partially  attacked  in  the  fusion  process. 

^  Use  5-6  cc.  of  an  acid  prepared  by  mixing  i  volume  of  concentrated  nitric 
acid  and  2  volumes  of  water. 


METALLIC   POISONS 


lf35 


upon  the  water-bath  with  dilute  hydrochloric  acid  (^12.5  per 
cent.)  and  potassium  chlorate  and  shake  frequently  or  stir. 
When  most  of  the  material  is  dissolved  and  the  solution  is 
yellow,  dilute  with  water.  Add  a  few  drops  of  sulphuric  acid 
and  filter  the  cold  solution. 


Material.     Treated  with  HCl  and  KCIO3.     Dilute  H2SO4.     Filter. 


Filtrate.^     Saturated  warm  with  H2S. 

Residue.      Tested 
for       "Metallic 

Precipitate.     Treated  with  hot 
(H4N)2Sx  and  (H4N)0H. 

Filtrate.      Tested 
for      "Metallic 
Poisons  III." 
Cr,  Zn. 

Poisons  IV." 
Ag,  Pb,  Ba. 

Filtrate.  Tested  for 
"Metallic      Poi- 
sons I." 

As,  Sb,  Sn,  Cu, 

Residue.     Tested 
for      "Metallic 
Poisons  II." 
Hg,  Pb,  Bi,  Cu, 
Cd. 

Action  of  Heavy  Metals 

Most  salts  of  heavy  metals,  as  lead,  copper,  mercury,  silver,  uranium  and 
bismuth,  precipitate  proteins.  These  compounds  are  metallic  salts  of  albumins 
(albuminates).  In  combining  with  the  oxides  of  these  metals,  proteins  behave 
like  acids.  If  the  metal  albuminates  first  formed  are  insoluble,  or  only  sUghtly 
soluble  in  the  body  fluids,  they  are  non-toxic  or  only  slightly  toxic.  But  a  sol- 
uble albuminate  is  transported  throughout  the  organism  and  exerts  a  toxic  action. 
Every  cell  in  contact  with  the  dissolved  metal  may  be  poisoned.  Mercury 
albuminate  is  an  example  of  the  latter  class  of  metal  albuminates.  Being  sol- 
uble in  sodium  chloride  and  protein  solutions,  it  acts  as  a  powerful  poison. 
Copper  albuminate  on  the  other  hand  is  not  appreciabty  soluble  in  solutions  of 
sodium  chloride,  hydrochloric  acid  or  proteins.  Not  entering  the  circulation,  it 
is  as  good  as  non-toxic.  Lead  and  silver  albuminates  are  like  copper  albuminate 
as  regards  solubility  in  the  solvents  mentioned.  But  if  a  heavj^  metal,  which 
forms  a  difficultly  soluble  albuminate,  finds  its  way  into  the  organism  in  organic 
combination  so  that  it  cannot  be  precipitated  by  proteins,  for  example,  copper  in 
union  with  tartaric  acid,  it  then  is  as  poisonous,  or  nearly  as  poisonous,  as 
mercury  in  corrosive  sublimate.  Administration  intravenously  of  20  mg. 
of  such  copper  causes  the  death  of  an  adult  rabbit. 


'  If  this  filtrate  contains  much  free  hydrochloric  acid,  remove  most  of  the 
acid  by  evaporation.  Then  add  ammonia  until  alkaline  and  finally  acidify 
with  dilute  nitric  acid. 


166  DETECTION   OE  POISONS 

Consequently  precipitation  takes  place  wherever  the  salt  of  a  heavy  metal 
comes  in  contact  with  proteins.  The  term  corrosion  is  applied  to  such  an 
occurrence.  There  is  always  present  the  metallic  oxide,  protein  and  the  acid 
originally  combined  with  the  metal.  As  a  rule  the  acid  is  loosely  held  by  the  pre- 
cipitate and  is  washed  away  by  the  circulating  blood.  The  corrosive  action  of 
salts  of  heavy  metals  is  due  both  to  the  union  of  the  metallic  oxide  with  protein, 
living  protein  being  changed  to  dead  metal  albuminate,  and  to  the  caustic  action 
of  the  free  acid.  Therefore  the  intensity  of  the  action  of  the  salt  of  the  heavy 
metal  depends  upon  the  nature  of  the  given  metal  albuminate.  The  degree  of 
solubihty  is  especially  important  (see  above) ,  also  the  quantity  and  strength  of  the 
free  acid.  Salts  of  heavy  metals  not  only  may  affect  the  place  of  apphcation  but 
they  may  give  rise  to  serious  changes  where  ehminated,  as  in  the  intestines  or 
kidneys.  Before  eHmination  they  may  also  seriously  harm  parenchymatous 
organs  Uke  the  liver,  as  well  as  the  circulatory  organs.  Finally  salts  of  heavy 
metals  have  an  important  action  upon  the  blood.  R.  Kobert  and  his  collabor- 
ators have  found  that  white  as  well  as  red  blood-corpuscles  may  combine  with 
metals  and  thus  act  as  antidotes.  Kobert  has  shown  that  the  substance  of  red 
blood-corpuscles  is  capable  of  taking  up  a  considerable  quantity  of  a  heavy  metal. 
A  chemical  compound  (metal  haemoglobin)  is  formed  and  the  oxyhaemoglobin 
spectrum  is  not  changed.  Thus  lead  speedily  impairs  the  vitality  of  red  blood- 
corpuscles.  Consequently  red  blood-corpuscles  are  killed  in  large  quantity  in 
lead  poisoning. 

Fate,  Distribution  and  Elimination  of  Metals  in  the  Human  Body 

Arsenic. — EUmination  of  arsenic  takes  place  mainly  through  the  urine.  It 
begins  several  hours  (7-12)  after  administration  and,  after  a  single  dose,  usually 
lasts  4-7  days.  A  great  many  experiments  have  shown  the  duration  of  arsenic 
elimination  in  urine  to  vary  from  a  few  days  to  several  weeks.  Some  observers 
have  found  arsenic  in  urine  80,  and  even  90  days,  after  poisoning.  Conse- 
quently in  suspected  arsenic  poisoning  first  examine  the  urine!  In  arsenic 
poisoning  the  urine  is  usually  diminished  in  volume  and  contains  albumin  and 
blood-corpuscles. 

As  regards  retention  of  arsenic  by  different  organs,  large  quantities  of  the  poison 
are  usually  found  in  the  liver.  Examine  also  the  stomach  and  intestines  with 
contents,  since  most  of  the  poison  will  obviously  be  in  these  organs  in  case  of 
recent  administration.  The  spleen,  kidneys  and  muscles  usually  contain  arsenic. 
But  the  brain  rarely  shows  more  than  traces  of  the  poison.  On  the  other  hand, 
the  bones  in  many  instances  have  contained  arsenic.  This  fact  has  given  rise  to 
the  hypothesis  that  arsenic  is  capable  of  replacing  phosphoric  acid  in  the  bones. 

During  the  first  stage  in  elimination  of  arsenic  from  the  organism  the  poison 
appears  to  be  deposited  in  the  bones  as  calcium  arseniate.  If  large  doses  of 
difficultly  soluble  arsenic  compounds,  as  white  arsenic,  or  small  doses  of  soluble 
compounds  have  been  taken,  the  liver  alone  appears  to  arrest  and  retain  the 
poison.     Probably  the  quite  stable  arseno-nucleins  are  formed  in  that  organ. 

From  experiments  where  the  organism  was  deluged  with  easily  soluble  arsenic 
compounds,  Chittenden  concluded  that  the  brain  can  arrest  and  retain  the  poison 


METALLIC   POISONS 


07 


and  that  arsenic  then  accumulates  there.  But  after  administration  of  white 
arsenic  or  Schweinturt  green,  only  traces  of  the  poison  can  be  detected  in  the 
brain.  Two  instances  of  arsenical  poisoning  are  in  favor  of  this  view.  In  the 
first  case  a  woman  died  9  hours  after  eating  a  highly  arsenical  soup.  The  liver 
weighing  1259  grams,  contained  76  mg.  of  arsenious  oxide;  the  kidneys  and 
bladder  0.6  mg.;  and  half  the  brain  only  a  faint  trace  of  the  poison.  In  the 
second  case  a  young  woman  died  a  day  after  taking  Schweinfurt  green.  In 
this  instance  too  the  brain  contained  only  traces  of  the  poison.  In  experiments 
upon  animals  most  of  the  arsenic  was  always  found  in  the  liver. 


Normal  Arsenic^ 

The  view  that  certain  organs  of  the  body  may  contain  arsenic  as  an  essential 
constituent  has  led  to  the  use  of  the  term  "normal"  to  distinguish  such  arsenic 
from  that  entering  the  organism  in  food  or  drugs.  The  introduction  of  this  term 
into  chemical  literature  is  unfortunate,  because  it  suggests  the  possibility  of  two 
kinds  of  arsenic.  Such  a  notion  has  no  foundation  whatever.  Arsenic  is  arsenic 
and  no  test  capable  of  showing  more  than  one  kind  is  known.  A  committee  of 
the  French  Academy  of  Sciences^  after  carefully  investigating  this  matter  came 
to  the  conclusion  that  arsenic  never  occurs  normally  in  the  human  body.  But 
within  recent  years  A.  Gautier^  after  making  many  analyses  of  different  materials 
has  come  to  the  opposite  opinion.  Gautier  thus  summarizes  his  results  in  one  of 
his  papers:^ 

"Speaking  from  a  medico-legal  point  of  view,  I  would  state  that  arsenic, 
aside  from  the  thyroid,  mammary  and  thymus  glands,  never  occurs  in  the  human 
body  except  in  the  skin,  hair,  bones,  milk  and  sometimes  in  the  faeces  and  then 
only  in  traces  which  are  often  infinitesimal.  Excepting  the  brain,  the  other  organs 
and  fluids,  especially  those  forming  the  bulk  of  the  body,  as  muscular  tissue,  liver, 
spleen,  kidneys,  lungs,  blood,  urine,  etc.,  fail  to  show  the  slightest  trace  of  arsemc 

If  a  chemist  therefore  examines  individually  these  arsenic- 
free  organs  by  my  method  or  by  one  less  delicate  and  finds  traces,  especially 
appreciable  traces,  of  this  metalloid,  such  arsenic  has  been  absorbed  during  life 
either  medicinally  or  criminally." 

Gautier  found  the  largest  quantity  of  arsenic  in  the  thyroid  gland  (0.75  mg. 
in  100  grams  of  gland)  but  this  result  has  not  yet  been  confirmed.  Other 
chemists  to  be  sure  have  found  arsenic  in  the  thyroid  gland  but  in  much  smaller 
quantity.  These  same  chemists  have  also  found  arsenic  in  organs  which  Gautier 
says  are  non-arsenical.  The  following  is  a  summary  of  their  results  but  the 
original  papers  should  be  consulted  for  full  details : 

1  The  brief  account  of  "normal  arsenic"  in  the  German  edition  seems  insuffi- 
cient. After  thoroughly  examining  the  literature,  the  translator  has  therefore 
decided  to  treat  this  subject  more  fully.     Tr. 

2  Comptes  rendus  de  I'Academie  des  Sciences  12,  1076-1109  (1S41). 

3  Ibid.,  129,  929-936  (1899). 
*Ibid.,  130,  284-291  (1900). 


168 


DETECTION   OP   POISONS 


Remarks  and 

Observer    ^ 

Material 

Arsenic  found 

conclusions 

Gautier^ 

Human  and  ani- 

o .75    mgr.    in    100 

Used  6  glands  and 

mal  organs  and 

grams   of   human 

assumed   uniform 

other  material 

thyroid  gland 

distribution  of  As 

Bertrand^ 

Only  animal  ma- 

0 .015  mgr.  per  100 

Concludes  As  is  a 

terial 

grams     of     dried 

normal    constitu- 

sponge 

ent  of  protoplasm 

Schaefer^ 

Human  organs 

0  .007  mgr.  per  100 

Concludes  As  may 

grams   of   human 

occur  in  all  organs 

thyroid 

but  found  many 
free  from  As 

Pagel^ 

Human  and  ani- 

Positive   but    not 

Found  testes  arseni- 

mal organs 

quantitative 

cal  but  Gautier 
says  they  are  not 

On  the  contrary  several  chemists  have  carefully  analyzed  human  and  animal 
organs,  either  finding  no  arsenic  or  detecting  this  metalloid  in  mere  traces,  which 
are  inconstant  in  occurrence  and  confined  to  no  special  organ.  The  table 
on  the  opposite  page   briefly  summarizes  their  results. 

The  results  set  forth  in  this  table  place  "normal  arsenic"  in  a  doubtful 
position  at  least.  If  it  is  a  reality  and  not  a  fancy,  the  quantity  of  arsenic, 
compared  with  that  obtained  in  an  analysis  actually  dealing  with  this  metalloid, 
is  so  minute  that  the  toxicologist  need  feel  no  concern.  If  he  has  conducted  his 
analysis  with  every  precaution  as  regards  reagents  and  method  and  obtained  a 
distinct  mirror,  he  may  dismiss  the  "normal  arsenic"  chimera  and  accept  the 
result  as  due  to  arsenic  that  has  entered  the  body  from  some  external  source. 
KunkeP  has  summed  up  the  matter  in  these  words. 

"The  so-called  normal  arsenic,  if  there  is  such  a  thing,  does  not  affect  the 
results  of  forensic  chemistry,  because  the  so-called  normal  quantities  are  so 
exceedingly  small  (o.oi  or  even  o.ooi  mg.  in  an  organ)  that  the  quantities 
necessary  to  furnish  a  satisfactory  forensic  proof,  which  are  a  hundred  or  even 
a  thousand  times  greater,  must  be  regarded  as  an  entirely  different  and  much 
higher  order  of  magnitudes." 

Antimony. — Elimination  of  antimony  takes  place  largely  through  the  urine. 
The  rest  of  the  metal  is  found  chiefly  in  the  liver  and  gastro-intestinal  tract,  as 
well  as  in  the  kidneys  and  brain.     Pouchet  found  antimony  in  bones,  skin,  hair 

^  Loc.  cit. 

^Annalesde  ITnstitutPasteur  17,1-10(1903);  Comptes  rendus  de  I'Academie 
des  Sciences  135,  809-812  (1902);  and  Bulletin  de  la  Societe  Chimique  de  Paris 
(3),  27,  1233-1236  (1902). 

^  Annales  de  Chimie  Analytique  12,  52-58  and  97-101  (1907). 

^  Dissertation,  University  of  Nancy,  1900. 

^  Zeitschrift  fiir  physiologische  Chemie  44,  511-529  (1905). 


METALLIC   POISONS 


109 


Observer 

Material 

Results 

Conclusion 

Hodlmoseri 

Human    thyroid 

20  analyses 

Liver  gives  positive 

gland  and  liver 

Negative    or    few 

results  as  often  as 

traces 

thyroid  gland 

Cerny2 

Human  and  ani- 

28 analyses 

Minute  traces  may 

mal  thyroid  and 

Negative  or  faint 

appear    but    not 

thymus  glands. 

traces 

constant 

Human  liver 

Ziemke^ 

Various     human 

Over    40  analyses. 

Not  a  normal  con- 

organs 

Negative.        One 

stituent  of  human 

case  doubtful 

organs 

Wieser'' 

Various     human 

32  analyses.  Mostly 

Traces    inconstant 

and  animal  or- 

negative   but    few 

and  due  to  chance 

gans 

traces 

contamination 

KunkeP 

Various     human 

Negative 

No  such  thing  as 

and  animal  or- 

normal As 

gans 

Bloemendal^ 

Various     human 

Mostly       negative 

No   such   thing   as 

and  animal  or- 

but few  traces 

normal  As 

gans 

Warren' 

Human    thyroid 

32  analyses.    Nega- 

No  such   thing   as 

gland 

tive     except     two 

normal  As 

- 

slight  traces 

and  chiefly  in  the  intestinal  tract.  A  large  part  of  the  ingested  poison  may  be 
eliminated  by  emesis.  A  part  is  apparently  retained  in  the  body  for  some  time, 
since  antimony  has  been  detected  in  the  liver  and  in  the  bones  months  after  the 
last  administration. 

Lead.— Lead  is  eliminated  in  urine  and  fasces.  Elimination  by  the  fasces 
always  exceeds  that  by  the  urine,  even  when  the  lead  has  not  been  taken  by 
the  mouth.  Mann^  (see  R.  Robert,  Intoxikationen) ,  for  example,  in  the  case 
of  two  patients  was  never  able  to  find  more  than  0.6  milligram  of  lead  in  the 
urine  collected  during  24  hours,  whereas  the  fffices  during  the  same  period  con- 
tained 2-3  milligrams  of  lead.  In  lead  poisoning  the  metal  has  been  found  in  the 
saliva,  bile  and  in  both  red  and  white  blood-corpuscles.     In  animals  relatively 

1  Zeitschrift  fiir  physiologische  Chemie  33,  329-344  (1901). 

-Ibid.,  34,  408-416  (1902). 

3  Vierteljahrsschrift  fiir  gerichtliche  Medizin  (3)  23,  51-60  (1902). 

^Dissertation,  University  of  Wiirzburg  (1903). 

*  Zeitschrift  fiir  physiologische  Chemie  44,  511-529  (1905). 

''Dissertation,  Universit}'  of  Leyden  (190S). 

'  W.  H.  Warren,  anah'ses  not  published. 

8  Zeitschrift  fiir  physiologische  Chemie  6,  6  (1882). 


170  '      DETECTION   OF   POISONS 

most  of  the  lead  has  been  found  in  the  kidneys,  after  which  come  bones,  liver, 
testes  and  finally  the  brain  and  blood.  In  experiments  with  sheep  Ulenberger 
and  Hofmeister  obtained  the  following  results: 

^  ,  r>   .  1  Pb  in  grams  per  looo 

Organs  and  iiuids 

grams 

Kidneys  o .  44-0 . 47 

Liver  0.3  -0.6 

Pancreas  o .  54 

Salivary  glands  0.42 

BUe  0.1 1-0.40 

Bones  0.32 

Faeces  0.22 

Spleen  0.14 

Blood  0.05-0.12 

Urine  o .  06-0 .  08 

Lead  is  eliminated  especially  by  the  bile  and  in  acute  poisoning  this  secretion 
may  contain  more  lead  than  any  of  the  other  organs  or  secretions.  Oliver^  has 
given  the  following  results  for  human  material: 

-.  Pb  in  grams  per  1000 

Organs 

grams 

Liver  0.0416 

Spleen  0.039 

Large  brain  0.0216 

Small  brain  o .  0086 

Kidneys  0.013 

Heart  o .  0005 

Elimination  of  lead  by  the  urine  is  said,  in  the  case  of  man,  not  always  to  be 
uniform.  The  urine  is  free  from  lead  for  a  long  time  and  later,  without  further 
administration,  again  contains  the  metal.  This  behavior  is  in  harmony  with  the 
fact  that  lead  can  be  retained  in  the  organism  many  months.  In  chronic  lead 
poisoning  the  brain  has  frequently  been  found  to  contain  much  lead.  In  this 
kind  of  poisoning  elimination  of  the  metal  is  always  more  abundant  by  the  faeces 
than  by  the  urine. 

Chromimn. — Chromic  acid  and  soluble  chromates  and  dichromates  are  quite 
toxic.  The  mucous  membranes  absorb  alkaline  chromates  rapidly  and  severe, 
acute  poisoning  occurs.  The  poison  causes  intense  pain  in  the  stomach  and 
intestines,  collapse  and  kidney  derangement  which  may  terminate  fatally  in  a 
few  hours.  Other  symptoms  are  nausea,  vomiting  of  yellow  matter  which  later 
is  tinged  with  blood,  diarrhoea  and  even  bloody  stools,  intense  thirst,  emaciation, 
great  anxiety,  severe  pain  in  the  abdomen,  faint  and  quickened  pulse — "the 
cholera  picture."  (Kunkel,  Toxikologie.)  The  statements  regarding  the  quan- 
tity of  an  alkaline  chromate,  capable  of  producing  acute  poisoning,  agree  fairly 
well.  Even  a  few  decigrams  (0.2  gram)  may  cause  very  serious  symptoms  which 
sometimes  terminate  fatally.     Chromic  acid  is  eliminated  mainly  by  the  urine  but 

^  The  Lancet,  March,  1891. 


METALLIC   POISONS  171 

partly  by  the  intestines.  Elimination  takes  place  rapidly  and  the  body  is  soon 
free  of  the  poison.  Four  days  after  administration  of  quite  large  quantities  of  a 
chromate,  the  urine  and  faices  are  said  to  contain  only  traces  of  this  metal. 

Copper. — Only  a  small  amount  of  copper,  varying  with  different  compounds,  is 
absorbed  by  the  intestines  and  carried  into  the  circulation.  Sodium  cupric 
tartrate  and  copper  salts  of  fatty  acids  are  absorbed  most  easily.  Copper  poison- 
ing rarely  occurs  from  introducing  a  copper  compound  into  the  stomach.  Copper 
compounds  in  large  amounts  act  as  local  caustics  and  occasion  severe  pain  in  the 
stomach.  Vomiting  and  the  sense  of  taste  make  it  impossible  to  take  much  of  a 
copper  compound.  Foods  containing  copper  are  unpalatable.  The  sense  of 
taste  as  well  as  after-taste  prevent  one  from  swallowing  such  food  in  any  quantity. 
Food,  containing  0.5  gram  of  copper  per  kilogram,  has  a  marked  taste.  Irresisti- 
ble nausea,  steadily  increasing,  soon  makes  it  impossible  to  take  more  of  the  food 
containing  copper.  Elimination  of  copper  by  the  urine  is  very  slight.  Copper 
absorbed  from  the  intestines  is  arrested  by  the  liver  where  it  accumulates. 
Traces  of  copper  have  frequently  been  found  in  the  human  liver.  The  author  in 
toxicological  analyses  has  repeatedly  found  weighable  quantities  of  copper  in 
the  livers  of  adults  who  had  not  taken  copper  salts  beforehand,  except  possibly  for 
suicidal  purposes.  The  liver  is  the  most  important  organ  for  the  detection  of 
copper,  next  to  which  come  the  bile,  kidneys  and  the  gastro-intestinal  mucosa. 
Copper  is  said  to  be  in  the  liver  as  a  nuclein  compound.  In  the  case  of  blood, 
copper  is  located  not  in  the  serum  but  in  the  corpuscles. 

Mercury. — Distribution  of  mercury  in  the  body  is  said  to  be  always  the  same, 
no  matter  what  the  method  of  administration  is.  It  is  immaterial  whether  it  is 
introduced  by  the  mouth,  hypodermically  or  from  an  abrasion.  Elimination  of 
mercury  takes  place  through  the  saliva,  sweat,  bile,  gastro-intestinal  mucosa  and 
urine.  Elimination  in  the  saliva  seems  to  be  constant,  since  mercury  can  always 
be  detected  in  the  saliva  during  use  of  mercurials  in  lues.  A  relatively  large 
quantity  of  this  metal  is  said  to  be  eliminated  in  the  sweat.  Opinions  differ  as  to 
the  relative  quantities  eliminated  by  the  urine  and  intestines.  Usually  elimina- 
tion by  the  intestines  exceeds  that  by  the  kidneys.  Recent  experiments  appear 
to  show  that  mercury  is  eliminated  in  the  urine  regularly  and  in  slowly  increasing 
quantity  and  then  slowly  diminishes.  Elimination  of  mercury  ceases  after  6-9 
months  and  even  later.  In  a  most  favorable  case  the  total  quantity  of  mercury 
eliminated  in  the  urine  amounts  to  about  50  per  cent,  of  the  total  quantity  taken 
but  is  frequently  much  less.  In  mercury  poisoning  the  kidneys,  of  all  the  organs 
persistently  contain  most  of  the  poison  even  for  weeks.  Then  follow  the  liver, 
spleen,  bile  and  intestinal  mucosa.  In  toxicological  analysis  the  urine  should 
also  be  examined,  though  in  acute  poisoning  it  always  contains  only  a  fraction  of  a 
milligram  of  mercury  in  a  Uter.  In  severe  mercurial  poisoning  the  metal  may  be 
said  to  occur  in  all  organs  and  secretions. 

Electrolytic  Separation  of  Mercury  from  Urine 

Heat  a  liter  of  urine  upon  the  water-bath  about  2  hours  vdth  5-6  grams  of 
potassium  chlorate  and  10  cc.  of  concentrated  hydrochloric  acid  and  shake  fre- 
quently. Evaporate  to  300  cc.  and  use  this  solution  for  the  electrolytic  separa- 
tion of  mercury.    Use  a  Bunsen  battery  of  3-4  cells,  or  any  other  galvanic  appara- 


172  DETECTION    OF   POISONS 

tus  having  the  same  strength  of  current.  A  thin  sheet  or  rod  of  gold  2  mm.  thick 
and  6-10  cm.  long  serves  as  the  cathode,  and  a  piece  of  platinum  wire  of  about  the 
same  thickness  as  the  anode.  Place  the  electrodes  in  the  solution  2-4  cm.  apart 
and  allow  the  experiment  to  run  24-48  hours.  Wash  and  dry  the  gold  cathode, 
upon  which  the  mercury  is  deposited,  and  place  in  a  glass  tube  (20  cm.  long  and  4-5 
cm.  in  diameter).  This  tube  is  sealed  at  the  bottom  and  reduced  at  the  top  to 
smaller  size.  Apply  heat  until  all  the  mercury  is  expelled  from  the  gold.  De- 
posit the  sublimate  3-4  cm.  beyond  the  top  of  the  gold  rod.  Then  seal  the  tube 
below  the  sublimate.  Introduce  a  small  crystal  of  iodine  into  the  tube  and  seal 
the  other  end.  Heat  the  iodine  carefully  over  a  small  flame  to  bring  it  into  con- 
tact with  the  mercury.     The  two  elements  combine  to  form  red  mercuric  iodide. 

Sometimes  it  is  convenient  to  precipitate  mercury  from  urine  upon  other 
metals,  for  example,  copper,  gold,  brass  and  zinc  dust.  Witz  heats  to  boiling 
with  hydrochloric  acid  and  concentrated  potassium  permanganate  solution  to 
destroy  organic  matter.  Use  10  cc.  of  concentrated  hydrochloric  acid  and  15-20 
cc.  of  potassium  perm_anganate  solution  for  500  cc.  of  urine.  Slowly  pass  the 
decolorized  liquid  through  a  glass  tube  over  a  copper  spiral.  Wash  the  copper 
with  potassium  hydroxide  solution,  then  with  absolute  alcohol  and  cleanse  with 
filter  paper.  Finally  dry  at  70-80°  and  heat  in  a  glass  tube.  Treat  the  mercury 
sublimate  with  iodine  as  described.  Satisfactory  deposition  of  mercury  upon 
other  metals  (gold,  copper)  requires  previous  destruction  of  organic  matter  in  the 
urine  by  hydrochloric  acid  and  potassium  chlorate.  Otherwise,  organic  substances 
deposited  upon  the  metal  interfere  with  the  iodine  test  for  mercury. 

Silver.- — In  severe  poisoning  silver  has  been  found  in  bile,  faeces  and  in  many 
organs.  In  acute  poisoning  the  urine  is  usually  free  from  silver,  whereas  the 
contents  of  the  intestines  may  contain  the  metal  even  after  subcutaneous  injec- 
tion. Absorbed  silver  salts  appear  to  be  reduced  in  all  parts  of  the  body.  Ex- 
amination of  the  bodies  of  persons  who  have  suffered  from  argyria  (chronic  silver 
poisoning)  has  disclosed  precipitation  of  metallic  silver  in  the  organs.  Silver 
salts,  added  to  albumin  solutions,  form  very  stable  compounds  usually  amor- 
phous. Silver  may  evince  even  greater  affinity  for  albumin  than  for  chlorine. 
All  observers  agree  that  only  very  little  of  the  silver  reaching  the  intestines  is 
absorbed.  After  silver  medication  the  stools  are  black  from  silver  sulphide. 
Frequent  quantitative  estimations  of  silver  in  argyrotic  organs  have  been  made. 
In  one  case  the  liver  gave  0.047  P^r  cent,  of  silver  and  the  kidneys  0.061  per  cent. 
In  the  condition  called  argyria,  or  argyrosis,  the  skin  is  black.  Internal  adminis- 
tration of  silver  salts  causes  this  color  to  develop  gradually.  By  degrees  it  may 
become  quite  marked  even  causing  disfigurement. 

Quantitative  Estimation  of  Silver  in  Organs  and  Urine 

V.  Lehmann,!  in  determining  silver  in  organs  (liver,  kidneys,  spleen,  brain), 
first  thoroughly  dries  the  finely  ground  material  and  then  fuses  with  sodium 
carbonate  and  potassium  nitrate.  Extract  the  melt  with  water  and  dissolve 
the  insoluble  residue  of  metallic  silver  in  hot  nitric  acid.  Evaporate  this  solution 
to  dryness  upon  the  water-bath  and  precipitate  silver  as  chloride.  Avoid  a 
large  excess  of  hydrochloric  acid. 

^  Zeitschrift  fiir  physiologische  Chemie  6,  19  (1882). 


METALLIC   POISONS  173 

Mix  urine  wilh  sodium  carbonate  and  potassium  nitrate,  evafiorate  to  dryness, 
fuse  the  residue  and  treat  the  melt  as  described. 

Uraniiim. — Experiments  made  by  R.  Kobert  have  shown  that  uranium, 
administered  subcutaneously  or  intravenously,  is  the  most  toxic  of  all  metals. 
Uranyl  acetate  is  an  excellent  precipitant  of  albumins  and  the  other  uranyl  salts 
must  behave  in  much  the  same  way.  Consequently  internal  administration 
of  concentrated  solutions  of  uranyl  salts  destroys  the  mucous  surfaces  they 
touch,  for  example,  that  of  the  stomach,  changing  the  living  stomach  wall  to 
dead  uranyl-albuminate.  Uranyl  salts  therefore  must  be  classed  among  the 
powerful  caustic  poisons.  In  addition  to  acting  as  local  corrosives,  uranium 
salts  resemble  hydrocyanic  acid  in  partially  arresting  internal  oxidation  in  the 
organs  and  occasioning  the  severest  disturbances  of  metabolism. 

Bismuth. — This  metal  becomes  quite  toxic  when  it  reaches  the  blood.  Bis- 
muth solutions,  prepared  by  dissolving  bismuthous  hydroxide  in  tartaric  or  citric 
acid  and  then  neutralizing  with  sodium  or  ammonium  hydroxide  solution,  have 
been  repeatedly  administered  to  animals  subcutaneously  and  intravenously. 
The  smallest  lethal  dose  of  these  double  bismuth  salts,  injected  subcutaneously, 
was  found  to  be  but  6  mg.  per  kilogram  for  a  dog  or  cat  and  24  mg.  per  kilo- 
gram for  a  rabbit.  Bismuth  salts  insoluble  in  water  produce  entirely  differ- 
ent results  when  administered  internally.  Bismuth  subnitrate  and  similar 
salts  dissolve  very  slightly  in  the  highly  diluted  hydrochloric  acid  of  the  gastric 
juice.  Consequently  very  little  bismuth  is  conveyed  to  the  blood.  Most  of  the 
bismuth  taken  by  the  mouth  reaches  the  intestines.  Instead  of  being  absorbed, 
it  is  changed  to  bismuthous  sulphide  by  the  hydrogen  sulphide  always  present. 
Absorbed  bismuth  is  eliminated  by  the  saliva,  bile,  urine,  mucous  lining  of  the 
mouth,  stomach,  small  and  large  intestine  and  also  the  milk.  If  an  animal  is 
poisoned  by  bismuth,  the  metal  can  be  detected  in  the  urine,  bile,  liver,  kidneys, 
spleen,  walls  of  the  intestines  as  well  as  in  the  bones.  Different  observ-ers  have 
found  the  metal  in  especially  large  amount  in  the  milk  but  very  little  in  the  kid- 
neys and  liver. 

Zinc. — There  is  no  doubt  that  zinc  salts  reaching  the  intestinal  tract  are 
absorbed  in  very  small  quantity.  As  yet  there  is  no  satisfactory  explanation  of 
the  fate  of  absorbed  zinc.  In  zinc  poisoning  large  amounts  of  the  metal  have  been 
found  repeatedly  in  the  liver  and  bile.  This  may  mean  that  zinc  is  arrested  by 
the  liver  and  eliminated  in  the  bile.  Lehmann^  after  335  days  killed  a  dog  that 
had  been  fed  for  a  considerable  time  upon  zinc  carbonate.  The  following  organs, 
arranged  according  to  the  quantity  of  metal  in  each,  contained  zinc:  liver,  bile, 
large  intestine,  thyroid  gland,  spleen,  pancreas,  urine,  kidneys,  bladder,  muscle, 
brain,  lymphatic  gland,  stomach,  small  intestine,  lungs,  blood.  Occasionally 
considerable  quantities  of  zinc  may  be  taken  %\ith  articles  of  food  and  drink. 
All  acids  dissolve  metallic  zinc  very  freely.  Even  water  containing  carbon  diox- 
ide is  a  solvent.  Consequently  it  may  be  in  drinking  water  from  galvanized 
pipes.  All  kinds  of  food  and  drink,  kept  in  zinc  vessels  or  vessels  coated  with 
zinc,  may  contain  more  or  less  of  this  metal.  Plants  grown  upon  soil  containing 
zinc  take  up  the  metal.  Zinc  has  also  been  found  repeatedly  in  parts  of  human 
cadavers  under  circumstances  precluding  all  possibility  of  poisoning  by  this 

^  Archiv  fiir  Hygiene  28,  291  (1896). 


174  DETECTION   OF   POISONS 

metal.  Even  considerable  quantities  of  zinc  have  been  found  in  the  human 
liver. 

Tin. — The  cases  of  tin  poisoning  thus  far  observed  resemble  those  of  copper 
and  zinc.  What  knowledge  there  is  regarding  the  toxic  action  of  absorbed  tin 
has  been  gained  from  experiments  upon  animals.  These  experiments  show  that 
small  quantities  of  tin  are  absorbed  and  eliminated  in  the  urine,  when  ordinary- 
tin  compounds  are  brought  into  the  stomach.  But  thus  far  distinct  symptoms 
of  poisoning  by  such  quantities  of  the  metal  have  not  been  confirmed.  (Kunkel, 
Toxikologie.) 

White^  failed  to  produce  poisoning  by  bringing  tin  into  a  dog's  stomach.  The 
animal  received  sodium  stannous  tartrate  in  increasing  doses  for  22  days,  the 
daily  amount  being  0.02-0.06  gram.  Yet  the  animal  absorbed  tin.  In  the  urine, 
during  an  experiment  lasting  8  days.  White  found  0.02  gram  of  tin.  But  the 
tin  salt  mentioned,  introduced  directly  into  the  circulation  of  the  animal,  was 
quite  toxic  in  its  action.  Stannous  chloride,  administered  for  a  very  long  time 
to  a  dog,  produced  symptoms  of  poisoning.  The  urine  in  this  case  contained 
small  quantities  of  tin.  Kunkel  (Toxikologie)  states  that  tin  has  a  very  slight 
poisonous  action.  Apparently  it  is  eliminated  very  rapidly  by  the  kidneys. 
Quite  probably  this  prevents  accumulation  of  the  metal  in  the  body  and  conse- 
quent poisoning.  The  fact  that  White  did  not  observe  toxic  symptoms,  after 
feeding  a  dog  for  22  days  with  relatively  large  quantities  of  easily  absorbable 
sodium  stannous  tartrate;  and  that  Ungar  and  Bodlander^  failed  to  produce 
derangements  with  the  same  compound,  until  it  had  been  administered  for  a  year, 
prove  that  tin  is  quite  free  from  toxic  properties.  Hence,  tin  vessels  may  be  used 
and  preserved  articles  of  food  containing  tin  have  practically  no  deleterious 
action  upon  health. 

1  Archiv  fiir  experimentelle  Pathologic  und  Pharmakologie  13,  53. 

2  Zeitschrift  fiir  Hygiene  2,  241. 


CHAPTER  IV 

POISONS  NOT  IN  THE  THREE  MAIN  GROUPS 

MINERAL  ACIDS 

Hydrochloric,  Nitric  and  Sulphuric  Acids 

To  detect  free  mineral  acid,  extract  a  portion  of  material  with 
cold  water,  filter  and  test  as  follows,  if  the  solution  is  strongly 
acid: 

1.  Methyl  Violet  Test. — Add  a  few  drops  of  an  aqueous 
(o.i  :iooo)  or  alcoholic  (i  :ioo)  solution  of  methyl  violet^  to 
a  small  portion  of  filtrate.  A  free  mineral  acid  produces  a  blue 
or  green  color. 

2.  Methyl  Orange  Test. — Add  a  few  drops  of  a  dilute  aqueous 
solution  of  methyl  orange^  to  the  filtrate.  A  red  color  indicates 
free  mineral  acid. 

3.  Congo  Paper  Test. — Even  very  dilute  solutions  of  free 
mineral  acids  turn  ''Congo  paper"  blue. 

4.  Giinzburg's  Test. — Mix  a  few  drops  of  the  filtrate  with 
3-4  drops  of  Giinzburg's  reagent^  and  evaporate  to  complete 
dryness  upon  the  water-bath,  or  over  a  small  flame.  Free 
hydrochloric  or  sulphuric  acid  gives  a  fine  red  or  reddish  yellow 
residue.     Nitric  acid  gives  more  of  a  yellowish  red  residue. 

1  Methyl  violet  is  the  hydrochloride  of  hexa-methyl-para-rosaniline: 
(CH3)2N.C6H4\  /CH=CH\         V 

>C  =  C<  >C  =  N(CH3)2C1  (quinoidal  f  orm) 

(CHs)2N.C6H/  \CH=CH/ 

or 

V 

(CH3)2N.C6H4^         /C6H4.  N(CH3)2C1 

(CH3)2N.C6H4/^    ^ ^ 

*  Methyl  orange  =  Dimethyl-amino-azobenzene-4-sulphonic  acid: 
4  I        r  4 

(CH3)2  N.C6H4.N  =  N.C6H4.S020H. 
The  sodium  salt  of  this  sulphonic  acid  also  appears  in  commerce  under  the  name 
"methyl  orange." 

'  See  page  314  for  the  preparation  of  this  reagent. 

175 


176  DETECTION    OF    POISONS 

5.  Sulphocyanate  Test. — Add  a  little  potassium  sulphocya- 
nate  solution  to  ferric  acetate  solution  and  dilute  with  water 
until  yellow.  Then  add  the  solution  to  be  tested.  Free  mineral 
acid  produces  a  blood  red  color.  Traces  of  free  mineral  acid, 
especially  if  considerably  diluted,  do  not  give  a  red  color  until 
several  minutes  have  elapsed. 

One  or  more  of  these  general  tests,  which  furnish  evidence  of  a 
free  mineral  acid,  must  always  accompany  the  special  tests  to 
be  described  later.  Not  only  free  mineral  acids  give  the  special 
tests  but  in  certain  cases  their  salts.  Chlorides,  sulphates  and 
nitrates  are  normal  constituents  of  nearly  all  vegetable  and 
animal  materials.  As  a  rule  an  examination  of  cadaveric 
material  for  mineral  acids  is  necessary  only  when  the  autopsy 
points  conclusively  to  such  poisoning.  That  is  to  say,  when 
there  are  characteristic  corrosions  and  discolorations  about  the 
face,  mouth,  oesophagus  and  stomach.  If  general  tests  show  the 
presence  of  free  mineral  acid,  make  special  tests  for  the  particu- 
lar acid. 

Hydrochloric  Acid 

1.  Chlorine  Test. — Warm  a  little  of  the  aqueous  extract,  if 
not  too  dilute,  with  finely  powdered  manganese  dioxide.  Free 
hydrochloric  acid  yields  chlorine,  recognized  by  its  color  and 
odor,  or  by  passing  the  gas  into  potassium  iodide  solution  and 
liberating  iodine.  Hydrochloric  acid  exclusively  does  not  give 
this  test.  A  chloride  (NaCl)  and  free  sulphuric  acid  give  chlor- 
ine under  the  same  conditions. 

2.  Distillation. — If  possible,  separate  hydrochloric  acid  from 
other  substances  by  distillation.  The  concentration  of  the 
acid  is  especially  important,  since  dilute  hydrochloric  acid  upon 
distillation  at  first  yields  only  water.  Hydrochloric  acid^ 
itself  does  not  begin  to  distil  until  the  concentration  is  about 
10  per  cent.  Since  a  dilute  hydrochloric  acid  is  usually  ex- 
amined, distil  the  material  mixed  with  water,  or  preferably  a 

1  In  the  distillation  of  100  cc.  of  i  per  cent,  hydrochloric  acid,  the  first  90  cc. 
of  distillate  will  contain  only  traces  of  hydrochloric  acid,  whereas  the  last  portion 
will  contain  most  of  the  acid. 


POISONS    NOT   IN    'JTJK    THREE    MAIN    GROUPS  177 

filtered  aqueous  extract,  nearly  to  dryness.  In  such  a  dis- 
tillation apply  heat  by  means  of  an  oil  bath.  To  detect  hydro- 
chloric acid  in  the  distillate,  acidify  with  dilute  nitric  acid  and 
add  silver  nitrate  solution.  Frequently  a  quantitative  estima- 
tion of  hydrochloric  acid  is  required.  In  the  absence  of  other 
acids,  titrate  the  distillate  with  o.i  n-potassium  hydroxide 
solution,  using  phenolphthalein  as  indicator.  Otherwise,  esti- 
mate the  acid  gravimetrically,  precipitating  with  silver  nitrate 
and  weighing  silver  chloride,  or  volumetrically  by  Volhard's 
method.  In  the  latter  case,  precipitate  hydrochloric  acid  with 
O.I  n-silver  nitrate  solution  in  excess  and  subsequently  estimate 
that  excess  by  titration  with  o.i  n- ammonium  sulphocyanate 
solution,  using  ferric  alum  as  indicator.  Since  the  human 
stomach  normally  contains  0.1-0.6  per  cent,  of  free  hydro- 
chloric acid,  an  examination  of  stomach  contents  for  this  acid 
must  always  include  a  quantitative  estimation. 

Nitric  Acid 

The  human  body  normally  contains  only  a  very  small  amount  of  nitrates. 
When  present,  they  are  due  usually  to  vegetable  foods  which  contain  small 
quantities  of  nitrates.  Human  urine  almost  always  shows  traces  of  the  salts  of 
nitric  and  nitrous  acids.  The  chemical  examination  of  cadaveric  material  need 
not  include  tests  for  nitric  acid,  unless  the  autopsy  affords  evidence  of  poisoning 
by  this  acid,  as  distinct  signs  of  corrosion  about  the  lips,  mouth,  oesophagus  and 
stomach  and  sometimes  perforation.  These  parts  are  more  or  less  yellow  or 
yellowish  brown.  A  yellow  froth  is  said  to  exude  from  the  mouth  and  nose  of 
the  cadaver.  Also  the  stomach  contents  are  yellow  in  concentrated  nitric  acid 
poisoning.  If  the  concentration  of  the  acid  is  less  than  20  per  cent.,  these  specifiic 
changes  may  not  appear  in  the  gastro-int6stinal  tract.  Nitric  acid  taken  inter- 
nally, dilute  or  concentrated,  appears  at  once  in  the  urine. 

Detection  of  Nitric  Acid 

I.  Distillation. — If  possible,  extract  the  material  direct  with 
water,  filter  and  test  the  filtrate  for  nitric  acid  in  the  usual  way. 
When  the  quantity  of  nitric  acid  is  large,  separate  it  from  other 
substances  by  distilling  the  filtered  aqueous  extract.  Apply 
heat  by  means  of  an  oil  bath.     Nitric  acid^  does  not  distil,  until 

^  If  100  cc.  of  I  per  cent,  nitric  acid  are  mixed  ^-ith  bread  crumbs  and  distilled, 
most  of  the  acid  will  be  in  the  final  10  cc.  of  distillate. 
12 


178  DETECTION   OF  POISONS 

it  reaches  a  definite  concentration.  At  the  same  time  a  large 
part  of  the  acid  combines  with  organic  substances,  if  any  are 
present,  forming  nitro-derivatives,  xanthoproteic  acid,  etc. 
Nitric  acid  may  also  cause  oxidation.  Consequently  the  dis- 
tillate does  not  contain  all  the  acid  originally  present.  The 
residue  from  such  a  distillation  is  usually  distinctly  yellow. 
Toward  the  end  of  distillation  brown  vapors  of  nitrogen  peroxide 
often  appear.  Such  a  distillate,  added  to  starch  paste  and  po- 
tassium iodide,  produces  an  immediate  blue  color  in  presence  of 
dilute  sulphuric  acid. 

To  detect  nitric  acid  in  the  distillate,  employ  the  following 
tests : 

2.  G.  Fleury's  Procedure. ^^ — Extract  the  finely  divided  mate- 
rial with  absolute  alcohol,  filter  and  add  slaked  lime  in  excess 
to  the  filtrate.  To  decompose  any  nitric  acid  ester  present,  let 
the  mixture  stand  12  hours,  filter  and  evaporate  the  filtrate  to 
dryness.  Dissolve  the  residue  in  95  per  cent,  alcohol,  expel 
alcohol  from  the  filtered  solution  and  finally  test  an  aqueous 
solution  of  the  residue  for  nitric  acid.  Fleury  has  obtained  by 
means  of  this  method  about  20  per  cent,  of  the  nitric  acid  from 
animal  material.  This  procedure  converts  the  acid  into  its 
calcium  salt  which  is  soluble  in  alcohol.  But  sodium  nitrate 
is  also  quite  soluble  in  95  per  cent,  alcohol  (1:50).  There- 
fore, if  the  final  residue  gives  a  faint  test  for  nitric  acid,  the  proof 
of  free  acid  in  the  original  material  is  not  conclusive.  The 
following  method  obviates  this  difficulty. 

3.  Baumert's  Procediire.^ — Neutralize  the  material  itself,  or 
its  aqueous  extract,  with  milk  of  lime,  dry  and  extract  with 
alcohol.  Or,  after  neutralization  with  milk  of  lime  or  calcium 
carbonate,  evaporate  to  a  syrup  and  mix  the  latter  while  stir- 
ring with  alcohol.  Distil  the  filtered  alcoholic  extract  obtained 
in  either  way,  dissolve  the  residue  in  water,  filter  and  evapo- 
rate the  solution.  Dissolve  the  residue  again  in  alcohol  and 
allow  this  solution  to  stand  for  several  hours  in  a  closed  flask 
with  about  the  same  volume  of  ether.     Filter  this  alcohol- 

^  Annales  de  Chimie  analytique  appliqu6e  6,  12. 
^Lehrbuch  der  gerichtlichen  Chemie,  second  edition  (1907). 


POISONS    NOT   IN    THE    THREE    MAIN    GROUPS  179 

ether  solution,  evaporate  the  solvent  and  dissolve  the  residue 
in  a  little  water.  Apply  the  following  nitric  acid  tests  to  this 
solution: 

(a)  Diphenylamine  and  Sulphuric  Acid  Test.^ — Blue  color. 
Add  a  few  drops  of  diphenylamine  suli)hate  solution'  to  the 

aqueous  extract,  or  distillate,  and  carefully  pour  this  mixture 
upon  pure  concentrated  sulphuric  acid  free  from  nitric  acid. 
If  nitric  acid  is  present,  a  blue  zone  appears  where  the  two  liquids 
meet. 

(b)  Brucine  and  Sulphuric  Acid  Test. — Red  color. 

Mix  the  liquid  to  be  tested  with  the  same  volume  of  brucine 
sulphate  solution^  and  carefully  pour  this  mixture  upon  pure 
concentrated  sulphuric  acid.  If  nitric  acid  is  present,  a  red 
zone  appears  where  the  two  liquids  meet. 

(c)  Ferrous  Sulphate  and  Sulphxxric  Acid  Test. — Saturate  the 
liquid  to  be  tested  with  pure  ferrous  sulphate  and  carefully 
pour  this  solution  upon  pure  concentrated  sulphuric  acid.  If 
nitric  acid  is  present,  a  black  zone  appears  where  the  two  liquids 
meet. 

(d)  Copper  Test. — Place  a  small  piece  of  clean  copper  (wire 
or  sheet)  in  nitric  acid  and  heat.  Red-brown  vapors  of  nitrogen 
peroxide  (NO2)  appear. 

Sulphuric  Acid 

Nearly  all  animal  and  vegetable  substances  normally  contain  sulphates.  Con- 
sequently an  examination  for  free  sulphuric  acid  must  exclude  its  salts.  There 
is  no  need  of  examining  cadaveric  material  for  the  free  acid,  unless  marked  corro- 
sion and  discoloration  of  lips,  mouth,  oesophagus  and  stomach  indicate  its  presence. 
There  are  eschars  upon  the  Hps  and  the  mucous  lining  of  the  mouth  is  gra}-ish 
white.  The  white  coating  on  the  back  of  the  tongue  may  have  been  dissolved 
exposing  the  firm,  brownish  muscular  tissue  beneath.     The  tongue  often  looks 

^Prepare  this  solution  by  dissolving  i  gram  of  diphenylamine,  (CsH5)2XH 
in  5  grams  of  dilute  sulphuric  acid  and  loo  cc.  of  water. 

2  Prepare  this  solution  by  dissolving  i  gram  of  brucine  in  5  grams  of  dilute 
sulphuric  acid  and  100  cc.  of  water.  The  sulphuric  acid  used  must  give  none  of 
the  tests  for  nitric  acid.  If  it  does  not  meet  this  requirement,  heat  in  a  platinum 
dish  to  expel  interfering  nitrous  substances.  Or  distil  the  acid  from  a  small 
retort,  rejecting  the  first  part  of  the  distillate. 


180  DETECTION   OF   POISONS 

as  if  it  had  been  boiled.  The  mucous  lining  of  the  oesophagus  is  much  wrinkled 
and  coated  gray;  Externally  the  stomach  is  usually  brown  or  slate-gray  and  its 
contents  black.  Frequently  in  sulphuric  acid  posioning  there  is  perforation  of 
the  stomach  wall  and  brownish  black  masses  find  their  way  into  the  abdominal 
cavity.  There  may  be  black  spots  in  the  stomach,  due  according  to  R.  Kobert 
(Intoxikationen)  not  to  charring,  as  previously  supposed,  but  to  brown-black 
hsematin.  Acids  decompose  the  blood-pigment  oxyhaemoglobin  mainly  into 
hsematin  and  protein  (globulin).  Methsemoglobin  and  hsematoporphyrin  may 
also  be  formed.  Acids  produce  the  latter  from  haematin  and  in  the  change  there 
is  loss  of  iron.  All  three  of  these  decomposition  products  of  the  red  blood-pig- 
ment, namely,  methsemoglobin,  haematin  and  haematoporphyrin  may  be  formed 
successively  and  then  appear  in  the  urine.  The  blood  in  the  stomach  walls  is 
often  acid  and  then  contains  chiefly  methsemoglobin  and  haematin.  The  mucosa 
of  the  intestines  even  far  down  may  be  grayish  white  and  strongly  acid. 


Detection  of  Siilphviric  Acid 

1.  Extract  the  finely  divided  material,  if  strongly  acid,  with 
cold  absolute  alcohol  and  after  some  time  filter.  The  solution 
contains  sulphuric  acid  but  not  sulphates.  Evaporate  the 
alcoholic  filtrate  upon  the  water-bath,  or,  if  the  volume  is  large, 
distil  the  alcohol.  Dissolve  the  residue  in  a  little  water  (lo  cc.) 
and  heat  the  solution  to  boiling  to  saponify^  ethyl  sulphuric 
acid.  Filter  and  test  the  filtrate  with  barium  chloride  or  lead 
acetate  solution.  To  prove  that  the  precipitate  is  a  sulphate, 
mix  with  sodium  carbonate  and  fuse  upon  charcoal.  The 
sodium  sulphide  formed  blackens  metallic  silver  in  presence  of 
water,  or  gives  hydrogen  sulphide  with  acids. 

2.  Extract  the  finely  divided  material  with  water  and  apply 
the  following  tests  to  the  filtrate: 

(a)  Sugar  Test. — Evaporate  some  of  the  filtered  extract  in  a 
porcelain  dish  with  a  small  particle  of  sugar.  Free  sulphuric 
acid  produces  a  black,  carbonaceous  residue. 

(b)  Sulphur  Dioxide  Test.- — Concentrate  the  filtered  extract 
upon  the  water  bath  and  heat  in  a  test-tube  with  a  few  pieces  of 
copper.  Free  sulphuric  acid  generates  sulphur  dioxide,  recog- 
nized by  its  stifling  odor.  Distil  the  sulphur  dioxide  (preferably 
in  an  atmosphere  of  carbon  dioxide)  into  a  little  water  and  test 
the  distillate  as  follows: 

1  HO.SO2.OC2HB  +  H2O  =  C2H5.OH  +  H2SO4. 


POISONS    NOT    IN    THE    'IIIKEK    MAIN    CKOV/PS       ■  181 

a.  Warm  some  of  the  liquid  with  a  Httle  stannous  chloride 
solution.     A  yellow  precipitate  of  stannic  sulphide^  appears. 

j8.  Add  iodo-potassium  iodide  solution  drop  by  drop.  The 
color  of  the  iodine  disappears  and  at  the  same  time  suljihuric 
acid  is  formed: 

H2SO3  +  H2O  +  I2  =  H2SO,  +  2HI. 

Barium  chloride  then  precipitates  barium  sulphate  insoluble 
in  dilute  hydrochloric  acid. 

To  estimate  sulphuric  acid  quantitatively,  either  precipitate 
and  weigh  barium  sulphate  in  the  usual  way,  or  titrate  with 
0.1  n-potassium  hydroxide  solution,  using  phenolphthalein  as 
indicator. 

1000  cc.  of  0.1  n-potassium  hydroxide  solution  =  o.i  gram- 
equivalent  of  sulphuric  acid  =  4.9  grams  of  H2SO4. 

Detection  of  Sulphvirous  Acid 

Sulphur  dioxide  acts  most  injuriously  when  inhaled.  It  is  very  irritating  to 
the  respiratory  organs  and  also  changes  the  blood-pigment.  After  death  the 
respiratory  organs  are  found  to  be  profoundly  altered  as  when  acted  upon  by 
strong  mineral  acids.  After  severe  poisoning  by  vapors  containing  sulphur 
dioxide,  the  blood  is  dirty  brownish  red^  and  usually  gives  the  hsematin  spectrum. 
Human  beings  experience  discomfort,  if  there  are  0.015-0.02  volumes  of  sulphur 
dioxide  per  1000  volumes  of  air.  Many  persons  become  quite  ill  in  a  few  minutes, 
when  there  are  0.03  volumes  of  sulphur  dioxide  in  1000  volumes  of  air.  The  gas 
produces  a  sharp,  stinging  sensation  in  the  nostrils,  sneezing  and  coughing.  In 
experiments  upon  mice,  rabbits  and  guinea-pigs,  Lehmann  observed  marked 
toxic  symptoms  from  air  containing  0.04  volume  per  cent,  of  sulphur  dioxide; 
death  ensued  in  6  hours  from  0.06  per  cent.;  and  in  20  minutes  from  0.08  per  cent. 
Articles  of  food  and  drink,  preserved  by  means  of  sulphurous  acid  or  its  salts, 
may  injure  the  health,  causing  especially  gastro-intestinal  catarrh  and  other 
chronic  derangements.  For  this  reason  it  is  prohibited  to  preserve  articles  of 
food  and  drink  by  means  of  sulphurous  acid,  sulphites  and  hj-posulphites. 

If  the' quantity  of  sulphur  dioxide  in  air  is  not  too  small,  its  presence  may  be 

^  Sulphurous  acid  and  sodium  sulphite,  added  to  stannous  chloride  solution 
not  too  strongly  acid,  precipitate  stannous  sulphite,  SnSOs,  white  and  readily 
soluble  in  hydrochloric  acid.  Warmed  in  presence  of  hj-drochloric  acid,  sulphur 
dioxide  acts  upon  a  stannous  salt  as  an  oxidizing  agent.  A  precipitate  of  SneOioSa 
is  formed,  or  H2S  is  evolved  and  SnCU  formed,  depending  upon  the  amount  of 
hydrochloric  acid  present.  (Prescott  and  Johnson,  Qualitative  Chemical  Analy- 
sis.    Fifth  edition,  page  86.) 

2  Neutral  sulphites  cause  the  blood  to  become  brick  red. 


182  DETECTION    OP   POISONS 

recognized  by  its  characteristic  stifling  odor.  A  strip  of  paper,  moistened  with 
a  solution  of  pure  potassium  iodate  (KIO3)  and  starch,  turns  blue  in  air  contain- 
ing sulphur  dioxide  owing  to  the  formation  of  a  compound  of  iodine  and  starch. 
This  reaction  serves  as  a  preliminary  test  for  the  detection  of  sulphurous  acid  and 
hyposulphites  in  chopped  meat,  sausage  meat  and  other  meat  products.  Shake 
the  meat  in  an  Erlenmeyer  flask  with  phosphoric  acid,  suspend  in  the  neck  of  the 
flask  from  the  stopper  (see  Fig.  i,  page  3)  a  paper  strip  prepared  as  described 
and  heat  the  flask  upon  the  water-bath.     The  paper  should  not  turn  blue. 

Explanation. — Sulphur  dioxide  reduces  potassium  iodate  (a).  Sulphuric  acid 
thus  formed  liberates  hydriodic  and  iodic  acids  from  their  salts  (/3  and7).  The 
iodine  set  free  by  the  interaction  of  these  two  acids  (5)  finally  turns  the  starch 
blue. 

(a)     KIO3  +  3H2SO3  =    KI      +  3H2SO4, 

(13)   2KI       -I-    H2SO4  =  2HI       +    K2SO4, 

(7)  2KIO3  +    H2SO4  =  2HIO3  +    K2SO4, 

(5)     HIO3  +  sHI        =  3I2        +  3H2O. 

The  official  directions^  for  the  detection  and  quantitative  estimation  of  sulphur 
dioxide  in  meat  are  as  follows.  Mix  30  grams  of  finely  chopped  meat  with  200 
cc.  of  boiled  water  in  a  500  cc.  distilling  flask.^  Add  sodium  carbonate  solution 
until  the  reaction  is  faintly  alkaline. 

Let  the  mixture  stand  for  an  hour  and  then  completely  expel  air  from  the  ap- 
paratus by  passing  carbon  dioxide  through  the  tube  extending  to  the  bottom  of 
the  flask.  Then  introduce  into  the  Peligot  tube  (see  below)  50  cc.  of  iodine 
solution  (5  grams  of  pure  iodine  and  7 . 5  grams  of  potassium  iodide  in  a  liter 
of  water) .  Raise  the  stopper  of  the  distilling  flask  and,  without  stopping  the  flow 
of  carbon  dioxide,  add  10  cc.  of  10  per  cent,  phosphoric  acid  solution.  Then 
carefully  heat  the  contents  of  the  flask  and  distil  half  the  liquid,  maintaining  all 
the  while  a  current  of  carbon  dioxide.  Transfer  the  contents  of  the  Peligot 
tube,  which  should  be  brown,  to  a  beaker,  rinsing  it  out  with  water  to  prevent 
loss  of  solution.  Add  a  little  hydrochloric  acid,  heat  and  by  means  of  barium 
chloride  solution  completely  precipitate  the  sulphuric  acid  formed  from  the  oxida- 
tion of  sulphurous  acid  by  iodine. 

H2SO3  +  H2O  +  I2  =  H2SO4  +  2HI. 

If  this  test  is  positive,  then  the  meat  examined  contains  either  free  sulphurous 
acid,  sulphites  or  hyposulphites.  In  the  quantitative  estimation  the  barium 
sulphate  should  be  weighed  in  the  usual  manner. 

OXALIC  ACID 

Oxalic  acid  and  its  salts,  for  example,  salt  of  sorrel,  are  quite  toxic  substances. 
Administration  of  oxalic  acid  has  terminated  fatally  in  the  case  of  adults  in  a  few 

1  Measures  for  putting  into  effect  the  law  of  the  German  Empire  of  June  3, 
1900,  relating  to  the  inspection  of  beef-cattle  and  meats. 

2  The  apparatus  prescribed  for  official  examinations  is  a  distilling  flask,  having 
a  capacity  of  400-500  cc.  and  provided  with  a  two-hole  stopper  for  two  glass 
tubes  entering  the  flask.  One  tube  extends  to  the  bottom  of  the  flask  and  the 
other  only  into  the  neck.  The  latter  is  connected  with  a  Liebig  condenser  to 
which  a  Peligot  tube  is  fastened  at  the  other  end  by  a  tight  stopper. 


POISONS    NOT   IN   THE    TIIKEE    MAIN    GROUPS 


183 


minutes.  Oxalic  acid  is  very  al)iin(lanl  in  the  vcf^cl able  kingdom  in  the  form  of 
its  acid  potassium  salt,  KIIC2()4,  and  calcium  salt.  Sorrel,  wood-sorrel  and 
rhubarb  are  especially  rich  in  salts  of  oxalic  acid.  Hence  this  acid  may  find 
access  to  the  body  through  food  and  drugs  of  vegetable  origin.  Moreover, 
oxalic  acid  is  a  normal  constituent  in  small  quantity  of  human  urine,  2-6  milli- 
grams being  excreted  in  the  course  of  a  day.  Consequently  in  examining  animal 
material  it  is  often  necessary  to  supplement  a  positive  qualitative  test  by  a 
quantitative  estimation  of  oxalic  acid. 

Toxic  Action. — An  important  difference  between  mineral  acids  and  oxalic 
acid  is  the  toxicity  of  salts  of  the  latter.  Not  only  do  free  oxalic  acid  and  its 
acid  potassium  salt,  salt  of  sorrel,  show  poisonous  properties  but  even  very  dilute 
solutions  of  neutral  sodium  oxalate,  Na2C204,  act  in  the  same  way.  Therefore 
in  oxalic  acid  poisoning  it  is  necessary  to  distinguish  between  local  corrosion, 
occurring  at  the  point  of  application  and  also  in  part  upon  elimination,  and  re- 
mote action  due  to  absorption.  Local  action  at  the  point  of  application  is  cor- 
rosive like  that  of  all  acids.  Local  action  at  the  place  of  elimination  depends  upon 
the  formation  and  insolubility  of  calcium  oxalate.  On  account  of  the  ease  with 
which  the  organism  takes  up  oxalic  acid  and  its  alkali  salts,  the  action  of  the  ab- 
sorbed poison  is  rapid.  The  effects  caused  by  its  presence  may  be  attributed 
to  the  fact  that  this  acid  removes  in  part  from  organs,  as  the  heart,  and  from 
body  fluids  (blood)  the  calcium  they  require  for  their  life  processes,  converting 
it  in  part  into  insoluble  calcium  oxalate.  Oxalates  diminish  the  coagulating 
power  as  well  as  the  alkalinity  of  blood.  On  the  other  hand  they  increase  the 
quantity  of  sugar  in  the  blood.  In  oxalic  acid  poisoning  there  is  a  depression 
of  the  entire  metabolism.  This  is  also  the  case  as  regards  taking  up  oxygen  and 
giving  off  carbon  dioxide.  The  body  temperature  falls  as  the  processes  of  metabo- 
lism are  retarded.  Owing  to  withdrawal  of  calcium  from  the  heart,  that  organ 
is  weakened  and  finally  paralyzed.  Local  action  upon  the  kidneys  is  due  to 
clogging  of  the  injured  urinary  tubules  by  deposits  of  calcium  oxalate.  The  flow 
of  urine  may  wholly  cease  in  consequence  of  total  impairment  of  the  urinary 
tubules  and  death  may  ensue  from  anuria  and  urasmia.  Fatal  poisonings  from 
large  doses  of  oxalic  acid  are  usually  of  short  duration.  R.  Robert  (Intoxika- 
tionen)  describes  a  case  where  death  occurred  within  10  minutes. 

Bischoffi  has  made  statements  in  regard  to  the  distribution  of  oxalic  acid  in 
the  different  organs  of  persons  poisoned  by  this  substance.  In  a  case,  which  ter- 
minated fatally  in  less  than  15  minutes,  the  quantity  of  oxalic  acid  in  each  organ 
was  determined  separately  and  found  to  be: 

Weight  Organ  Oxalic  Acid 

2240  grams  Stomach,    oesophagus,    intestine    and 

contents  2.28      grams. 

770  grams  Liver  0.285    grams. 

290  grams  Ridneys  0.0145  grams. 

180  grams  Blood  from  the  heart  0.0435  grams. 

40  grams  Urine  0.0076  grams. 


^Berichte  der  Deutschen  chemischen  Gesellschaft,  16,  1350  (1883). 


184 


DETECTION   OF  POISONS 


The  quantity  of  oxalic  acid  in  the  liver  is  noticeably  large.  The  kidneys  and 
urine  contain  only  a  little  of  the  poison,  owing  to  the  short  duration  of  life  after 
poisoning.  A  striking  thing  about  the  urine  excreted  during  oxalic  acid  poisoning 
is  the  abundant  deposition  of  crystallized  calcium  oxalate. 


Detection  of  Oxalic  Acid 

To  detect  oxalate  without  discriminating  between  the  free 
acid,  acid  potassium  salt  or  calcium  oxalate,  employ  the  follow- 
ing method: 

Add  to  the  finely  divided  material  3-4  volumes  of  alcohol 
and  acidify  with  dilute  hydrochloric  acid.  Stir  frequently  and 
let  the  mixture  digest  1-2  hours  cold.  Then  filter  through  a 
plaited  paper  moistened  with  alcohol  and  wash  the  residue  with 
alcohol.  To  prevent  formation  of  ethyl  oxalate  during  evapo- 
ration, add  about  20  cc.  of  water  to  the  total  filtrate.  Evapo- 
rate upon  the  water-bath  until  all  alcohol  is  expelled.  Pass 
the  aqueous  residue  through  a  small  filter.  Extract  the  filtrate 
in  a  separating  funnel  3-4  times  with  5c»-6o  cc.  portions  of  ether. 

Let  the  total  ether  extract  stand  for 
some  time  in  a  dry  flask,  then  pass 
through  a  dry  filter  and  distil.  Dis- 
solve the  residue  in  2-3  cc.  of  water 
and  pass  the  solution,  if  necessary, 
through  a  moist  filter.  Add  am- 
monium hydroxide  solution  until 
alkaline  and  then  saturated  calcium 
sulphate  solution.  If  there  is  a  pre- 
cipitate, acidify  with  acetic  acid  and 
let  solution  and  precipitate  stand 
over  night  in  a  covered  beaker.  If 
there  is  still  a  crystalline  precipitate,  it  can  be  only  calcium 
oxalate.  A  microscopic  examination  of  this  precipitate  is 
advisable.  Calcium  oxalate  forms  characteristic  octahedrons 
having  the  so-called  envelope  shape  (Fig.  17).  When  thor- 
oughly washed,  calcium  oxalate  may  be  converted  by  ignition 
into  calcium  oxide  which  may  be  weighed. 

CaO  :  H2C2O4.2H2O  =  Weight     of     CaO  :  x 
(56)  (126)  found 


Fig.  17. — Calcium  Oxalate 
Crystals. 


POISONS    NOT   IN    THE    THREE    MAIN    GROUPS  185 

Calculation. — Since  the  quotient  5O  :  126  =  0.444,  multiply  the  weight  of 
calcium  oxide  found  by  0.444  to  get  the  corresponding  amount  of  crystaHized 
oxalic  acid. 


FREE  ALKALIES 

Potassium,  Sodium  and  Ammonium  Hydroxides 

Free  Alkalies. — The  same  general  principles  used  in  detecting  mineral  acids 
are  applicable  also  to  the  alkalies.  Since  potassium  and  sodium  compounds  are 
normal  constituents  of  animal  and  vegetable  organisms,  and  since  ammonia  is  a 
decomposition  product  of  nitrogenous  organic  matter,  the  examination  must 
always  show  that  the  alkalies  are  in  the  free  state,  for  they  alone  and  their  car- 
bonic acid  salts  decompose  and  corrode  animal  tissues  and  not  their  neutral  salts. 

Poisonings  due  to  caustic  alkalies  resemble  those  caused  by  corrosive  acids. 
If  taken  internally,  their  corrosive  action  gives  rise  to  pain  in  the  mouth,  throat, 
oesophagus,  stomach  and  abdomen.  Mineral  acid  corrosions  are  dry  and  brittle, 
whereas  those  from  caustic  alkalies  are  soft  and  greasy.  The  alkali  albuminates 
formed  become  gelatinous,  swell  and  may  partly  dissolve  in  presence  of  much 
water.  The  destructive  action  of  the  caustic  alkalies  extends  deep  and  affects 
the  parts  around  the  corroded  places.  In  caustic  alkaline  solutions  gelatinous 
tissues,  horny  substances,  hair  and  skin  swell  considerably  and  finally  dissolve. 
The  stomach  in  alkali  poisoning  is  softened  in  places,  corroded  and  decidedly 
bright  red  in  color. 

Detection  of  Alkalies 
Ammonia 

Free  ammonia  is  usually  recognized  by  its  odor.  A  piece  of 
moist  red  litmus  paper,  held  over  the  material,  becomes  blue. 
A  paper  moistened  with  mercurous  nitrate  solution  is  blackened. 

Distillation. — If  the  material  is  strongly  alkahne,  extract 
several  times  with  absolute  alcohol.  Use  a  flask  with  a  glass 
stopper  and  distil  the  combined  extracts.  Collect  the  distillate 
in  a  little  dilute  hydrochloric  acid  and  evaporate  the  solution 
in  a  porcelain  dish  to  dryness  upon  the  water-bath.  Dissolve 
the  residue  in  water  and  test  the  solution  for  ammonia,  using 
Nessler's  reagent  and  chloroplatinic  acid. 

Fixed  Alkalies 

The  residue  from  the  above  distillation  may  contain  potas- 
sium and  sodium  hydroxides.  If  the  residue  is  strongly  alka- 
line, first  add  a  few  drops  of  phenolphthalein  and  then  excess  of 


186  DETECTION   OF   POISONS 

barium  chloride  solution.  The  red  color  and  the  alkahne  reac- 
tion, if  due  to  carbonates,  disappear,  because  two  neutral  salts 
are  formed: 

K2CO3  +  BaCl2  =  BaCOs  +  2KCI. 

But  if  alkaline  hydroxides  are  present,  the  alkaline  reaction 
and  red  color  remain,  for  soluble  barium  hydroxide  is  formed: 

2KOH  +  BaCl2  =  Ba(0H)2  +  2KCI. 

And  the  solution  of  this  compound  reddens  phenolphthalein. 

To  distinguish  potassium  from  sodium  hydroxide,  neutralize 
the  residue  from  distillation  with  dilute  hydrochloric  acid  and 
test  for  potassium  and  sodium  as  follows : 

1.  Add  solution  of  chloroplatinic  acid  (H2PtCl6)  which  causes 
the  precipitation  of  potassium  in  the  form  of  the  double  chloride 
of  potassium  and  platinum  (potassium  chloroplatinate,  K2PtCl6). 

2.  Add  de  Konink's  reagent^  which  is  a  solution  of  sodium 
cobaltic  nitrite,  6NaN02.Co2(N02) 6-  This  reagent  produces  a 
yellow  precipitate  of  potassium  cobaltic  nitrite,  6KNO2. 
Co2(N02)6  +  XH2O,  in  a  solution  containing  a  potassium  salt. 
To  hasten  the  reaction,  add  a  few  drops  of  acetic  acid. 

3.  Test  for  sodium  in  a  neutral  solution  by  adding  a  few  drops 
of  freshly  prepared  acid  potassium  pyro-antimonate  solution, 
K2H2Sb207.  At  first  the  solution  is  turbid  but,  if  stirred,  de- 
posits a  white  crystalline  precipitate  of  sodium  pyro-antimon- 
ate, Na2H2Sb207. 

VitaH's  procedure  in  testing  for  caustic  alkalies  consists  in 
shaking  the  alcoholic  extract  of  the  material,  prepared  as  far  as 
possible  with  exclusion  of  air  (see  above) ,  with  freshly  precipi- 
tated and  well-washed  mercurous  chloride.  Free  alkali  black- 
ens this  compound.  The  solubility  of  mercurous  oxide  (Hg20), 
the  black  compound  formed,  in  dilute  nitric  acid  distinguishes 
it  from  mercuric  sulphide. 

Quantitative  Estimation  of  Hydroxides  and  Carbonates  of 
Alkalies. — To  determine  both  the  free  caustic  alkali  and  that 

^  Prepare  sodium  cobaltic  nitrite  by  dissolving  10  grams  of  pure  sodium  nitrite 
and  4  grams  of  cobaltous  nitrate  separately  in  sufi&cient  water.  Mix  the  solutions, 
add  2  cc.  of  acetic  acid  and  dilute  to  100  cc.  with  water. 


POISONS  NOT  IN  THE  THREE  MAIN  GROUPS       187 

converted  into  carbonate,  first  determine  total  alkalinity  by 
titrating  a  portion  of  the  distillation  residue  with  normal  or 
O.I  n-hydrochloric  acid,  using  methyl  orange  as  indicator. 
Then  precipitate  carbonate  in  a  second  portion  of  the  distilla- 
tion residue  with  barium  chloride  solution  and  determine  free 
caustic  alkali  in  the  filtrate.  If  the  examination  shows  only 
alkaline  carbonate,  this  does  not  exclude  the  possibihty  of  caus- 
tic alkali  having  been  originally  present. 

POTASSIUM  CHLORATE 

Toxic  Action. — Large  doses  (4-10  grams)  of  polassium  chlorate,  KCIO3,  are 
decidedly  toxic.  During  the  first  stage  of  intoxication,  alteration  in  the  shape  of 
the  red  corpuscles  and  conversion  of  oxyhaemoglobin  in  the  intact  corpuscles  into 
brown  methaemoglobin  take  place.  Then  the  red  blood  corpuscles,  at  least  in  a 
case  of  severe  poisoning,  change  their  form,  becoming  shriveled  and  undergoing 
decomposition.  Toxicologists  (see  R.  Kobert,  Intoxikationen)  ascribe  change  of 
blood  pigment  and  red  blood  corpuscles  to  specific  salt  action  possessed  in  high 
degree  by  potassium  chlorate.  This  explanation  also  accounts  for  salt  diuresis, ^ 
appearing  at  the  beginning  of  potassium  chlorate  poisoning,  whereby  the  blood  is 
much  thickened.  But  most  notable  is  the  high  alkalinity  of  the  urine,  resulting 
in  decreased  alkalinity  of  the  blood  plasma.  In  severe  chlorate  poisoning  so  much 
oxyhfemoglobin  is  changed  to  methaemoglobin  that  the  amount  of  oxygen  in 
the  blood  may  drop  to  i  per  cent.  As  a  result  human  beings  or  animals  thus 
poisoned  may  become  asphyxiated  from  lack  of  oxygen.  Potassium  chlorate 
through  the  action  of  potassium  weakens  the  heart.  In  chlorate  poisoning  the 
blood  has  a  characteristic  chocolate-brown  color  (see  above). 

Potassium  chlorate  taken  by  the  mouth  is  quite  rapidly  eliminated  by  the 
kidneys.  After  administration  of  o.i  gram  of  potassium  chlorate,  chloric  acid 
appears  in  the  urine  in  an  hour.  Most  of  the  potassium  chlorate  passes  into  the 
urine  unchanged,  only  a  little  of  the  salt  being  reduced  to  potassium  chloride. 
During  chlorate  poisoning,  the  urine  is  usually  very  dark,  even  black,  and  may 
contain  hsemoglobin  and  methaemoglobin.  It  is  frequently  opaque  and  strongly 
alkaline.  Upon  long  standing  a  dark  brown  sediment  gradually  deposits.  The 
urine  also  contains  considerable  albumin. 

In  suspected  chlorate  poisoning,  the  urine  should  if  possible  receive  a  thorough 
chemical  and  microscopical  examination.  An  anuria  lasting  several  daj's  maj- 
precede  death  and  render  an  examination  of  the  urine  quite  impossible. 

Detection  of  Chloric  Acid 

To  isolate  potassium  chlorate  from  organic  material,  use  a 
dialyzer  which  should  be  as  flat  as  possible,  because  the  thinner 
the  layer  in  the  inner  container  and  the  larger  the  volume  of 

^  Diuresis  =  increased  secretion  of  urine. 


188  DETECTION   OP   POISONS 

water  in  the  outer  vessel,  the  more  rapid  the  diffusion.  Place 
the  material'  to  be  examined,  as  parts  of  organs  and  stomach 
or  intestinal  contents,  in  the  inner  container  of  a  flat  dialyzer 
and  pure  water  in  the  outer  vessel.  Allow  dialysis  to  take 
place  5-6  hours  without  changing  the  water  in  the  outer  vessel. 
Then  evaporate  the  dialysate  (contents  of  the  outer  vessel)  to 
dryness  in  a  porcelain  dish  upon  the  water-bath.  Dissolve  the 
residue  in  a  little  water  and  examine  the  filtered  solution  for 
chloric  acid  as  follows: 

1.  Indigo  Test. — Add  dilute  sulphuric  acid  and  a  few  drops 
of  indigo  solution,  until  there  is  a  distinct  blue  color.  Then 
introduce  sulphurous  acid  drop  by  drop.  If  chloric  acid  is 
present,  the  blue  color  changes  to  yellow  or  greenish  yellow. 
This  is  a  delicate  test  for  chloric  acid,  given  even  by  0.0 1  gram 
of  potassium  chlorate. 

2.  Silver  Nitrate  Test. — Add  silver  nitrate  solution  in  excess. 
If  there  is  a  precipitate  (AgCl),  filter  and  add  a  few  drops  of 
sulphurous  acid  to  the  clear  filtrate.  A  chlorate  will  cause  the 
precipitation  of  more  silver  chloride.  Silver  chloride  differs 
from  silver  sulphite  in  being  insoluble  in  hot  dilute  nitric  acid. 
Sulphurous  acid  reduces  silver  chlorate  to  chloride: 

AgClOs  +  3H2SO3  =  AgCl  +  3H2SO4. 

3.  Free  Chlorine  Test. — A  solution  containing  a  chlorate, 
heated  with  concentrated  hydrochloric  acid,  gives  free  chlorine. 
The  gas  passed  into  potassium  iodide  solution  liberates  iodine. 
Shake  the  solution  with  chloroform  which  dissolves  iodine  with 
a  violet  color.  This  test  indicates  chloric  acid  only  in  the  ab- 
sence of  substances  like  chromic  acid  and  dichromates  which 
also  give  chlorine  with  hydrochloric  acid. 

If  the  material  is  a  powder,  dissolve  in  water  and  filter  if 
necessary.  A  direct  test  for  chloric  acid  is  usually  possible  with 
such  a  solution. 

Quantitative  Estimation  of  Chloric  Acid 

To  estimate  potassium  chlorate  quantitatively  in  urine,  dialy- 
sates  and  other  liquids,  reduce  with  zinc  dust,  or  employ 
Scholtz's  method. 


POISONS    NOT    JN    THE    THREE    MAIN    GROUPS  189 

1.  Zinc  Dust  Method. — Divide  the  solution  into  two  equal 
parts.  Determine  chloride  gravimetrically  in  one  portion  by 
precipitating  and  weighing  AgCl,  or  volumetrically  by  titrating 
according  to  Volhard's  method. 

Determine  chloride  and  chlorate  together  in  the  second 
portion.  Add  5-10  grams  of  zinc  dust  and  a  little  dilute  sul- 
phuric or  acetic  acid,  and  heat  the  mixture  0.5-1  hour  upon  a 
boiling  water-bath.  Filter  and  wash  the  residue  with  boiling 
water.  Acidify  the  filtrate  with  nitric  acid  and  precipitate 
chloride  with  silver  nitrate.  More  chlorine  appears  in  the  sec- 
ond than  in  the  first  determination.  Calculate  the  percentage 
of  potassium  chlorate  from  the  difference  between  the  two 
chlorine  determinations.  One  molecule  of  KCIO3  upon  re- 
duction yields  i  molecule  of  KCl  and  therefore  i  atom  of  chlo- 
rine. 

Zinc  dust  in  presence  of  sulphuric  or  acetic  acid  reduces  potassium  chlorate  to 
chloride : 

(a)   KCIO3  +  3Zn  =  KCl  -|-  3ZnO, 

(^)  ZnO      4-  2CH3.COOH  =  H2O  +    Zn(CH3.COO)2. 

2.  Method  of  M.  Scholtz.^ — This  method  makes  use  of  the 
reducing  action  of  nitrous  acid  upon  chloric  acid: 

HCIO3  +  3HNO2  =  HCl  +  3HNO3. 

Add  to  the  solution  10  cc.  of  nitric  acid  (sp.  gr.  1.2  =  32  per 
cent.)  and  10  cc.  of  10  per  cent,  sodium  nitrite  solution.  Let 
the  mixture  stand  for  15  minutes  at  room  temperature.  Then 
add  30-50  cc.  of  0.1  n-silver  nitrate  solution  and  5  cc.  of  satu- 
rated iron  alum  solution,  (H4N)2S04.Fe2(S04)3.24H20.  Ti- 
trate excess  of  silver  with  o.i  n-ammonium  sulphocyanate 
solution.  1000  cc.  of  o.i  n-AgNOs  =  0.1  KCIO3  gram  = 
12.245  grams  of  KCIO3. 

The  slight  excess  of  nitrous  acid  has  no  effect  upon  the  deli- 
cacy of  the  reaction.  Liquids  Kke  dialysates  of  stomach  con- 
tents and  organs  always  contain  chloride.     In  that  case  first 

^  Archiv  der  Pharmazie  243,  353  (1905). 


190  DETECTION   OF  POISONS 

determine  the  amount  of  chloride  in  another  portion  by  Vol- 
hard's  method. 

H.  Hildebrandt^  has  adapted  Scholtz's  method  to  the  ex- 
amination of  urine.  First  completely  precipitate  chloride  in  a 
measured  volume  of  urine  with  silver  nitrate  in  presence  of 
nitric  acid.  Add  sodium  nitrite  solution  to  the  clear,  chloride- 
free  filtrate,  as  well  as  more  silver  nitrate  solution,  until  there 
is  no  longer  a  precipitate.  Determine  as  usual  the  weight  of 
silver  chloride  obtained. 

In  the  case  of  urine  a  larger  quantity  of  nitrous  acid  is  decomposed  by  the  urea: 
CO(NH2)2  +  2HNO2  =  CO2  +  2N2  +  2H2O. 
Consequently  do  not  use  too  little  sodium  nitrite. 

Behavior  of  Potassium  Chlorate  in  Putrefaction 

C.  Bischoff  states  that  potassium  chlorate,  mixed  with  moist, 
organic  substances,  especially  blood,  is  very  soon  reduced  to 
chloride!  Bischoff  describes  several  cases,  in  which  poisoning 
by  potassium  chlorate  had  undoubtedly  occurred,  and  yet 
chloric  acid  could  not  be  detected  chemically  in  parts  of  the 
cadaver. 

In  an  experiment,  100  grams  of  blood,  0.5  gram  of  potassium 
chlorate  and  100  grams  of  water  were  allowed  to  stand  for  5 
days  at  room  temperatures.  Not  a  trace  of  chloric  acid  could 
be  detected  in  the  dialysate.  Bischoff  concludes  from  this 
experiment  that  potassium  chlorate,  mixed  with  moist  organic 
substances,  especially  with  blood,  is  soon  reduced.  Conse- 
quently, chloric  acid  may  not  be  detected,  even  in  cases  of 
rapidly  fatal  poisoning  by  potassium  chlorate. 

Detection  of  Chlorate  in  Meat 

The  German  law  of  June  3,  1900,  relating  to  the  inspection  of  beef-cattle  and 
meat,  forbids  the  use  of  chlorates  in  preserving  meat,  sausage  and  fat.  The 
official  directions  prescribed  for  the  chemical  examination  of  meat  and  fats  are  as 
follows : 

Let  30  grams  of  finely  divided  meat  stand  i  hour  in  the  cold  with  100  cc.  of 
water  and  then  heat  to  boiling.     Filter  when  cold  and  add  silver  nitrate  solution 

^  Vierteljahrsschrift  fiir  gerichtliche  Medizin  32,  81  (1906). 


POISONS    NOT   IN   THE    THEEE    MAIN   GROUPS  Hil 

in  excess  to  the  filtrate.  Add  2  cc.  of  10  per  cent,  sodium  sulphite  solution  and 
2  cc.  of  concentrated  nitric  acid  to  50  cc.  of  the  clear  filtrate  from  the  silver  pre- 
cipitate and  then  heat  to  boiling.  If  there  is  a  precipitate,  insoluble  in  more  hot 
water  and  consisting  of  silver  chloride,  chlorate  is  present. 

SANTONIN,  SULPHONAL  AND  TRIONAL 

These  substances  do  not  find  a  place  in  the  Stas-Otto  process 
on  account  of  their  behavior  toward  cold  tartaric  acid  solution 
and   ether.     Use   the   following   method   for   their   detection. 

Extract  the  material,  neutral  or  faintly  acid  with  tartaric 
acid,  under  a  reflux  condenser  with  boihng  absolute  alcohol. 
Filter  hot  and  evaporate  the  filtrate  to  dryness  upon  the  water- 
bath.  Dissolve  the  residue  in  hot  water.  If  the  solution  is 
colored,  digest  for  some  time  upon  the  water-bath  with  bone- 
black  and  stir  frequently.  Filter  the  hot  solution.  All  of  the 
above  substances,  if  present  in  considerable  quantity,  crystalUze 
in  part  as  the  solution  cools.  Extract  the  filtrate  and  any 
crystals  thoroughly  with  chloroform  several  times.  Pass  the 
chloroform  extract  through  a  dry  filter.  The  residue  from 
chloroform  may  contain  santonin,  sulphonal  and  trional,  as 
well  as  acetanilide  and  phenacetine. 

The  chloroform  residue  may  also  contain  those  substances 
extracted  in  the  Stas-Otto  process  from  the  acid  solution  by 
ether.  Chloroform  completely  extracts  substances  like  anti- 
pyrine,  caffeine,  acetanilide,  phenacetine  and  salicylic  acid. 
As  a  rule  they  are  purer  from  this  solvent  than  from  ether. 
The  chloroform  residue  may  also  contain  the  weak  base  narco- 
tine. 

SANTONIN 

Santonin,  CisHisOs,  crystallizes  in  colorless,  inodorous,  shining  leaflets  which 
are  bitter  and  melt  at  170°.  Santonin  dissolves  in  5000  parts  of  cold  and  250 
parts  of  boiling  water;  in  44  parts  of  alcohol;  and  in  4  parts  of  chloroform.  All 
these  solutions  are  neutral.  It  is  slightly  soluble  in  ether  (i  :  150).  Light  turns 
these  crystals  yellow.  Upon  evaporation,  an  alcoholic  solution  of  the  yellow 
modification  deposits  white  santonin. 

Constitution.— Santonin  is  the  internal  anhydride  (lactone)  of  santonic  acid, 
CibH2o04.  Caustic  alkalies,  as  well  as  calcium  and  barium  hydroxides,  dissolve 
santonin  forming  salts  of  this  acid.  In  this  case,  as  with  aU  lactones,  the  lactone 
ring  is  broken  as  follows: 


192 


HaC 

I 

oc 


DETECTION   OE 

POISONS 

CHs 

CH3 

1       H2 

i       H2 

c    c 

c    c 

C      CH- 

-0\ H 

>C0  +  0K  = 

H2C      C      CH— OH 

1        1 

1        1        1 

C      CH- 

OC      C       CH— CH- 

v/\/ 

1 

\/\/           1 

c    c 

CH3 

C      C            CHs 

1    H2 

1      H2 

CH3 

CH3 

Santonin 

Potassium  santonate 

A  solution  of  a  santonate,  acidified  with  hydrochloric  acid,  first  gives  free 
santonic  acid.  To  isolate  this  compound  from  the  mixture,  extract  at  once  with 
ether.  Otherwise,  the  acid  loses  i  molecule  of  water  upon  standing  and  passes 
into  its  interna]  anhydride,  santonin. 

Santonin  is  also  a  ketone.  As  such  it  forms  a  hydrazone,  C1BH18O2  =  N  - 
NH.CeHs,  with  phenylhydrazine  and  an  oxime,  Ci6His02  =  NOH,  with  hy- 
droxy lamine. 

According  to  the  structural  formula  above,  santonin  is  a  derivative  of  hexa- 
hydro-dimethyl-naphthalene.  Fused  with  potassium  hydroxide,  santonic 
acid  gives  hydrogen,  propionic  acid  and  a  naphthalene  derivative,  namely, 
dimethyl-;8-naphthol. 

Behavior  in  the  Organism. — Santonin  seems  to  be  incompletely  absorbed  in  the 
body.  M.  Jaffe^  has  administered  quite  large  quantities  of  santonin  to  dogs  and 
rabbits.  He  obtained  a  new  substance,  called  a-oxysantonin  (C15H18O4),  from 
the  urine  of  the  dog,  amounting  to  5  or  6  per  cent,  of  the  santonin  administered. 
He  extracted  with  chloroform  considerable  quantities  of  unaltered  santonin  from 
the  faeces  of  the  dog.  Rabbits  can  usually  tolerate  being  fed  with  santonin  for 
weeks,  and  a-oxysantonin  is  formed  only  in  very  small  quantity.  In  the  ether 
extract  of  the  rabbit's  urine,  Jaffe  found  a  second  santonin  derivative,  /3-oxy- 
santonin,  isomeric  with  a-oxysantonin,  with  considerable  unaltered  santonin.  In 
these  experiments  only  about  half  the  santonin  administered  was  absorbed  by  the 
rabbit. 

After  administration  of  santonin,  a  red  pigment  called  santonin  red  appears  in 
human  urine.  Even  after  medicinal  doses  santonin  urine  is  red,  or  becomes  at 
least  scarlet-red  to  purple  on  addition  of  potassium  or  sodium  hydroxide  solution. 
Urine  containing  santonin  also  becomes  carmine  red  on  addition  of  calcium  hy- 
droxide solution. 

Detection  of  Santonin 

Ether,  benzene,  or  better  chloroform,  extract  santonin  only 
from  acid  solutions.  The  organic  solvent  fails  to  remove  this 
compound  from  an  alkaline  solution,  as  it  is  then  in  the  form 
of  a  santonate.     Santonin  is  not  an  alkaloid  and  forms  no  pre- 

^  Zeitschrif t  fiir  physiologische  Chemie  22,  537  (1896-1897). 


POISONS    NOT   IN    THE    TIIKKE    MAIN    GROUPS  103 

cipitates  with   the  general   alkaloidal  reagents,   but  it  gives 
several  more  or  less  characteristic  color  reactions. 

1.  Alcoholic  Potassium  Hydroxide  Test. — Pure  santonin, 
heated  with  an  alcoholic  solution  of  potassium  hydroxide,  gives 
a  fine  carmine  red  color,  which  gradually  changes  to  reddish 
yellow  and  finally  fades  entirely.  In  this  test  yellow  santonin 
dissolves  at  once  with  a  yellowish  red  color. 

2.  Sulphuric  Acid-Ferric  Chloride  Test. — Heat  santonin  with 
concentrated  sulphuric  acid  and  add  a  drop  of  ferric  chloride 
solution.  The  mixture  becomes  violet.  Use  about  i  cc.  of 
sulphuric  acid  to  o.oi  gram  of  santonin. 

3.  Furfurol-Sulphuric  Acid  Test. — Mix  2-3  drops  of  alcohoUc 
santonin  solution  with  1-2  drops  of  2  per  cent.  alcohoHc  furfurol 
solution  and  2  cc.  of  pure  concentrated  sulphuric  acid.  Warm 
this  mixture  in  a  small  porcelain  dish  upon  the  water-bath. 
A  purple-red  color  appears  and  changes  with  continued  heating 
to  crimson-red,  blue-violet  and  finally  to  dark  blue  (Thater^) . 

Only  a  few  alkaloids  and  glucosides  give  distinct  color  reactions  with  furfurol 
and  sulphuric  acid.  Substances  behaving  similarly  are  veratrine,  picrotoxin 
(violet)  and  piperine  (green  to  blue-green,  finally  indigo-blue).  The  colors  given 
by  a-  and  /3-naphthol  with  furfurol  and  sulphuric  acid  are  also  characteristic. 

SULPHONAL 

Sulplional,  C7H16O4S2,  crystalUzes  in  colorless,  inodorous  and  tasteless  prisms, 

melting  at  125-126°  and  distilling  with  slight  decomposition  at  300°.     It  is  soluble 

p-TT  in  500  parts  of  cold  and  15  parts  of  boiling  water;  in  135 

I  parts  of  ether;  and  in  65  parts  of  cold  and  2  parts  of 

CH3 — C — SO2.C2H6       boiling  alcohol.      Sulphonal  is  freely  soluble  in  chloro- 

1  form.       Especially  characteristic  of  this  compound  are 

""    ^    ^  the  ease  with  which  it  crystallizes  and  its  great  stabiUty 

in  presence  of  chemical   reagents.     The  halogens,  halogen  acids,  alkaUne  h)'- 

droxides  and  carbonates,  concentrated  sulphuric  and  nitric  acids  are  without 

action  in  the  cold. 

Preparation. — The  condensation  of  ethyl  mercaptan  (2 
molecules)  with  acetone  (i  molecule)  by  means  of  dry  hydrogen 
chloride  gas,  or  concentrated  sulphuric  acid,  results  in  the  forma- 
tion of  the  ethyl-mercaptole  of  acetone.  The  latter  compound, 
shaken  with  a  saturated  solution  of  potassium  permanganate 

^  Archiv  der  Pharmazie  235,  410  (1S97). 
13 


194  DETECTION   OF   POISONS 

in  presence  of  dilute  sulphuric"  acid,  undergoes  oxidation  with 
formation  of  sulphonal:^ 

HsCs  HSC2H6  HsCs        /SC2H6  +  O2  HaCv        /SO2C2H5 

)C=0+  =H20+  >C<  >  >C<; 

H3C/  HSC2H5  H3C/      \SC2H5  +  O2  H3C/     \SO2C2H5 

Acetone  Ethyl  Ethyl-mercaptole  Sulphonal 

mercaptan  of  acetone 

Detection  of  Stilphonal 

Ether,  or  better  chloroform,  extracts  sulphonal  from  acid, 
neutral  and  alkaHne  solutions.  Test  the  residue  left  upon 
evaporating  these  solutions  as  follows: 

1.  Melting-point  Test. — Determine  the  melting  point  (125- 
126°)  of  the  perfectly  pure  substance.  Crystallization  from 
boiKng  water  with  the  use  of  a  little  bone-black  easily  gives  a 
pure  product.  A  mixture  of  these  crystals  with  pure  sul- 
phonal should  also  melt  at  125-126°. 

2.  Reduction  Test. — Sulphonal  heated  in  a  test-tube  with 
powdered  wood  charcoal  gives  the  characteristic  odor  of  ethyl 
mercaptan. 

3.  Detection  of  Sulphur. — (a)  With  Sodium.  Fusion  of 
sulphonal  in  a  dry  test-tube  with  a  little  metalHc  sodium  pro- 
duces sodium  sulphide.  Dissolve  cautiously  (unaltered  metallic 
sodium!)  the  cold  melt  in  a  little  water,  filter  and  test  the 
filtrate  with  sodium  nitroprusside  solution  for  sulphide  (see- 
page 23). 

(b)  With  Potassium  Cyanide. — Fuse  i  part  of  sulphonal 
and  about  2  parts  of  pure  potassium  cyanide  in  a  dry  test- 
tube.  Note  the  penetrating  odor  of  ethyl  mercaptan  (C2H6.SH) . 
Potassium  sulphocyanate  is  also  a  product  of  the  reaction.  .  An 
aqueous  solution  of  the  melt,  acidified  with  dilute  hydrochloric 
acid,  becomes  deep  red  with  1-2  drops  of  ferric  chloride  solution. 

(c)  With  Powdered  Iron. — Sulphonal  heated  with  pure  pow- 
dered iron  free  from  sulphur  gives  a  garlic-like  odor.     Add 

^  Sulphur  in  the  sulphone  group  =  SO2  is  most  Hkely  sexivalent,  corresponding 
to  the  atomic  grouping  I,  and  not  quadrivalent,  as  in  II: 
VI  ^O  IV  /O 

I.   =sf  ;  11.   =S<(  I 


POISONS    NOT   IN    THE    THREE   MAIN   GROUPS  195 

hydrochloric  acid  to  the  residue.     Hydrogen  sulphide  evolved 
blackens  lead  acetate  paper. 

Detection  of  Sulphonal  in  Urine 

Sulphonal  is  cumulative  in  its  action.  Therefore  continuous  administration 
for  a  long  time  of  large  doses  may  result  in  the  collection  of  a  considerable 
quantity  of  this  compound  in  the  organism.  Most  of  the  sulphonal  taken  ap- 
pears in  the  urine  as  ethyl-sulphonic  acid,  C2H6-SO2OH.1  The  formation  of 
this  acid  causes  an  increase  of  ammonia  in  the  urine  during  sulphonal  intoxication, 
as  does  administration  of  mineral  acids. 

Sulphonal  occurs  in  urine  in  detectable  quantity  only  following  considerable 
doses,  especially  when  they  have  been  taken  without  interruption.  Such  urine 
is  often  dark  red  to  garnet-brown  from  haematoporphyrin.  But  this  decom- 
position product  of  blood  pigment  appears  in  urine  only  succeeding  severe 
poisoning  by  sulphonal,  and  even  then  its  occurrence  is  rare. 

To  isolate  sulphonal  from  urine,  evaporate  1000  cc.  to  one-tenth  its  volume, 
and  extract  several  times  with  large  quantities  of  ether.  Pass  the  ether  extracts, 
after  they  have  settled  in  a  dry  flask  for  several  hours,  through  a  dry  filter  and 
distil.  Evaporate  the  residue  with  20  -30  cc.  of  10  per  cent,  sodium  hydroxide 
solution  to  dryness  upon  the  water-bath.  This  will  remove  coloring  matter, 
extracted  from  urine  by  ether,  but  will  not  affect  the  sulphonal.  Extract  sul- 
phonal from  the  alkaline  residue  with  ether.  Evaporate  the  solvent,  and  sul- 
phonal wiU  remain  pure  and  almost  colorless.  Determine  the  melting  point  of 
this  residue,  and  make  the  other  tests  for  sulphonal. 

Detection  of  Haematoporphyrin  in  Urine 

Coloring  matters  have  been  observed  in  red,  brownish  red  to  cherr\--red 
urines,  which  quite  probably  are  identical  with  haematoporphyrin.  The 
spectroscopic  examination  of  such  urine  is  made  in  the  following  manner.  Add 
sodium  hydroxide  solution,  drop  by  drop,  to  about  500  cc.  of  urine,  until  the 
reaction  is  strongly  alkaline,  and  then  add  a  Uttle  barium  chloride  solution. 
Filter  after  a  while,  and  wash  the  precipitate  well.  Extract  the  precipitate 
upon  the  filter  with  hot  alcohol,  containing  a  few  drops  of  dilute  sulphuric  acid. 
A  spectroscopic  examination  of  this  filtrate  can  be  made  directly  mth  a  Brown- 
ing pocket  spectroscope.  Acid  haematoporphyrin  solutions  are  violet;  when 
more  concentrated,  they  have  a  cherry-red  color,  and  show  the  characteristic 
spectrum  with  two  absorption  bands  (see  page  299).  If  the  acid,  alcoholic 
solution  is  saturated  with  a  few  drops  of  ammonium  or  sodium  hydroxide 
solution,  the  spectrum  of  alkahne  hasmatoporphjTin  solution  with  its  four  ab- 
sorption bands  appears.     Traces  of   haematoporphyrin  very  frequently  appear 

/° 
^  The  structural  formula  of  ethyl-sulphonic  acid  is:  C2H5.S;^OH. 

/O 
It  should  not  be  confused  with  ethvl-sulphuric  acid:    CoHi-O-Ss— OH. 

^O 


196  DETECTION   OF   POISONS 

in  normal  urine.     It  has  been  observed  more  abundantly,  at  times,  in  urine 
during  chronic  sulphonal  poisoning. 

TRIONAL 

Trional  crystallizes  in  colorless,  shining  leaflets  melting  at  76°.     It  is  soluble 

in  320  parts  of  cold,  but  more  easily  soluble  in  hot  water.     It  is  also  soluble  in 

alcohol,  ether  and  chloroform.     The  aqueous  solution  is 

^■^3\^     /^^2-C2Hb     neutral  and  bitter.     In  the  latter  respect  it  differs  from 

P  jj  /    ^SO   C  H       sulphonal  which  is  tasteless.     Trional  gives  the  sulphonal 

reactions.      Trional    is   completely   decomposed  in  the 

organism  and  the  danger  of  cumulative  action  is  much  less  than  in  the  case  of 

sulphonal.     Moreover,  haematoporphyrin  has  almost  never  been  observed,  even 

following  considerable  doses  of  trional  and  after  uninterrupted  use  for  weeks. 

Active  Organic  Substances^  Rarely  Occurring  in  Toxicological 
Analysis 

CANTHARIDIN 

Cantharidin,   C10H12O4,   is   the    active    vesicating    principle   of   Spanish  fly 

(Lytta  vesicatoria)  and  is  present  to  the  extent  of  0.8-1  per  cent.     Cantharidin 

TT  forms    colorless,    shining,   neutral,   rhombic  leaflets, 

C         CH2.COOH  melting  at  218°  and  subliming  at  higher  temperature 

/  I  \/'  in   white   needles.     It  is   almost  insoluble   even  in 

H2C     I     C     O  boiling  water.     Acids,  as  tartaric  acid,  increase  its 

s^^  solubility  in  water,  though  cantharidin  is  not  a  base. 

jj  r;         Q CO  It  dissolves  with  difiiculty  in  cold  alcohol  (0.03 :  100  at 

\    /  18°)  and  in  ether  (o.i 1 :  100).    Chloroform  (1.52  :  100), 

C  acetone  and  acetic  ether  are  its  best  solvents.     It  is 

TT 

^  as  good  as  insoluble  in  petroleum  benzine. 

Constitution. — According  to  H.  Meyer^  cantharidin  has  the  structural  formula 
shown  above.  This  compound  is  at  the  same  time  a  monobasic  acid  and  a 
/3-lactone.  Potassium  or  sodium  hydroxide  breaks  the  labile  /3-lactone  ring. 
Cantharidin  passes  into  solution  as  the  alkali  salt  of  diabasic  cantharidic  acid, 
C10H14O5: 

H  K    OH  H 

C  CH2— COO    ;H  C 

/  I  \    /  /  I  \     /CH2— COOK 


H2C          C— O  H2C  I      C<^ 

I    CH2I       I  =         I  CH2I   ^OH                        +H2O. 

\      ...  ± .H..  (       \| 

H2C         C— CO                OK  H2C          C— COOK 

\    /  \    / 

c  c 

H2  H2 

Cantharidin  Potassium  cantharidate 

1  The  toxic  substances  considered  in  this  place  have  been  arranged  in  alpha 
betical  order. 

^Monatshefte  fiir  Chemie  18,  393  (1897)  and  19,  707  (1898). 


POISONS    NOT   IN   THE   THREE    MAIN   GROUPS  1-37 

Potassium  cantharidatc,  C^om20,K,.2lU),  rccenLly  rccommcmlecl  for 
phthisis,  and  sodium  cantharidatc,  CoHisOBNa^aHjO,  are  well  crystall>zed 
salts  Mineral  acid  first  sets  cantharidic  acid  free  from  these  salts.  The  latter 
soon  loses  a  molecule  of  water,  passing  into  its  internal  anhydride,  canthandin. 


H 


H 


C  CHa— COOH  C  CHj— COOH 

'l\      /  /    \    / 

H  H2C    I      C— O 


H2C     I      C— 0 
I    CH2I 


HoC 


CH2I       I  +  H2O 

C— CO  iOH  H2C         C— CO 


Cantharidic  acid  Cantharidin 

Cantharidin,  heated  with  hydriodic  acid  at  100°,  or  treated  at  room  tem- 
perature with  chlorosulphonic  acid,  CI-SO2-OH,  changes  into  the  isomeric 
cantharic  acid,  C10H12O4,  crystallizing  in  colorless  needles  melting  at  275  . 
This  add  is  not  a  vesicant.  Heated  for  3  hours  at  135°  in  sealed  tube  with 
acetyl  chloride,  cantharic  acid  yields  another  isomer  of  cantharidin  caUed  iso- 
cantharidin  (Anderlini  and  Ghiro).i  The  latter  crystallizes  from  alcohol  in 
large  colorless  leaflets  melting  at  76°.  There  is  a  close  relationship  between  0- 
xylene  and  cantharidin,  for  the  latter,  heated  at  400°  with  calcium  hydroxide, 
gives  a  dihydro-o-xylene,  CsHis,  called  cantharene,  and  also  o-xylene,  CeHr 
(CH3)2,  and  xyHc  acid.  Finally,  cantharidin,  heated  with  an  excess  of  phos- 
phorus pentasulphide  and  distilled,  gives  pure  o-xylene.     (J.  Piccard.)^ 

Detection  of  Cantharidin 

Evaporate  a  liquid,  or  material  containing  much  moisture 
(organs,  stomach  or  intestinal  contents,  etc.),  to  dryness  upon 
the  water-bath.  Dragendorff  directs  repeated  extraction  of  the 
finely  divided  material  with  alcohol  containing  sulphuric  acid. 
Filter  the  extracts,  add  one-sixth  their  volume  of  water  and 
distil  the  alcohol.  Extract  the  residue  2-3  times  with  chloro- 
form and  shake  the  chloroform  extracts  with  water  to  remove 
adherent  acid.  Finally  separate  the  chloroform^  from  water, 
distil  and  examine  the  residue  for  cantharidin.  Since  this  com- 
pound gives  no  characteristic  chemical  reactions,  employ  the 
physiological  test  for  identification.  Dissolve  the  chloroform^ 
residue,  unless  fatty  substances  are  present,  in  a  few  drops  of 

1  Berichte  der  Deutschen  chemischen  Gesellschaft  24,  199S  (1S91). 

2  Ibid.,  i2,S77  (1879)- 


198  DETECTION   OE  POISONS 

hot  almond  oil.  Bind  a  cloth,  saturated  with  this  solution, 
upon  the  upper  arm  or  breast  by  means  of  adhesive  plaster. 
Cantharidin  reddens  the  skin  and  sometimes  raises  blisters. 
Even  0.14  mg.  of  cantharidin  causes  blistering.  Salts  of  can- 
tharidic  acid  also  have  a  vesicating  action. 

To  detect  cantharidin  in  blood,  brain,  liver  and  other  material 
rich  in  protein,  E.  Schmidt  boils  with  dilute  potassium  hydroxide 
solution  (i  gram  of  KOH  in  15  cc.  of  water),  until  the  mass  is 
homogeneous,  acidifies  with  dilute  sulphuric  acid  and  extracts 
thoroughly  with  hot  alcohol.  The  procedure  in  other  respects 
is  as  described  above. 

Cantharidin  is  said  to  resist  putrefaction. 

CYTISINE 

Cytisine,  C11H14N2O,  occurs  in  the  ripe  seeds  of  Golden  chain  (Cytisus  Labur- 
num) which  contain  about  1.5  per  cent.  Cytisine  and  the  alkaloid  originally 
called  ulexine,  isolated  from  the  seeds  of  Ulex  europaeus,  are  identical  (A. 
Partheil). 

Preparation. — Extract  powdered  ripe  laburnum  seeds  with  60  per  cent,  alcohol 
containing  acetic  acid.  Distil  the  alcohol  from  the  extracts,  pour  the  residue 
through  a  moist  filter  and  precipitate  extractive  and  tannin  substances  with 
lead  acetate  solution.  Filter,  add  potassium  hydroxide  solution  to  the  clear 
filtrate  and  extract  cytisine  with  chloroform.  Distil  the  chloroform  which 
usually  deposits  cytisine  as  a  radiating  crystalline  mass.  If  purification  is  neces- 
sary, recrystallize  the  residue  from  absolute  alcohol  or  boiling  hgroin.  Sub- 
limation in  a  partial  vacuum  also  purifies  crude  cytisine. 

Cytisine  crystallizes  in  large,  colorless,  tasteless  prisms,  melting  at  152-153° 
and  subliming  at  a  higher  temperature,  if  carefully  heated.  It  dissolves  freely 
in  water,  alcohol,  chloroform  and  acetic  ether;  less  easily  in  commercial  ether, 
benzene  and  acetone;  and  is  insoluble  in  petroleum  ether  and  absolute  ether. 

Cytisine  is  a  strong  secondary  base  and  very  toxic.  Although  capable  of  com- 
bining with  I  or  2  molecules  of  hydrochloric  acid,  this  compound  behaves  in 
other  respects  like  a  monacid  base.  Only  the  salts  containing  one  equivalent 
of  acid  crystallize  well.  Nitrous  acid  converts  this  secondary  base  into  nitroso- 
cytisine,  C11H13ON-NO,  which  crystallizes  in  needles.  Nitrous  fumes  appear, 
if  cytisine  is  warmed  upon  the  water-bath  with  twice  the  amount  of  concentrated 
nitric  acid,  and  the  solution  at  once  becomes  reddish  yellow  to  brown.  This 
solution  poured  into  water  gives  a  precipitate  of  nitro-nitroso-cytisine,  C11H12ON- 
(N02)N-N0.  This  compound  crystallizes  from  water  in  pale  yellow  scales 
melting  at  242-244°. 

Toxic  Action. — Cytisine  produces  convulsions,  its  action  in  this  respect  being 
very  similar  to  that  of  strychnine.  But  unlike  the  latter  alkaloid  it  also  irri- 
tates the  gastro-intestinal  mucosa  even  causing  bloody  inflammation.     Cytisine 


POISONS   NOT  IN   THE   THREE   MAIN   GROUPS  199 

also  differs  from  strychnine  in  stimulating  the  vomiting  center.  Consequently, 
after  doses  of  cytlsine  or  laburnum  preparations,  human  beings  and  animals 
capable  of  emesis  thus  rid  the  organism  of  a  large  part  of  the  poison.  Like 
strychnine  cytlsine  stimulates  the  respiratory  and  vaso-motor  centers.  Finally 
as  in  strychnine  intoxication  death  results  from  paralysis  of  these  two  centers 
A  part  of  the  cytisine  leaves  the  organism  unchanged  and  appears  in  the  urine 
(R.  Kobert.) 

Detection  of  Cytisine 

Prepare  an  aqueous  tartaric  acid  solution  of  stomach  con- 
tents, vomitus  or  parts  of  organs,  following  the  general  pro- 
cedure for  alkaloids.  To  remove  final  traces  of  fatty  acids  and 
fat,  shake  this  solution  well  with  ether.  Withdraw  the  aqueous 
solution,  make  alkaline  with  sodiurn  hydroxide  solution  and 
extract  thoroughly  with  chloroform  or  isobutyl  alcohol.  Evap- 
orate the  chloroform  or  isobutyl  alcohol  extracts  and  test  the 
residue  as  follows  for  cytisine : 

1.  Van  der  Moer's^  Test. — Ferric  chloride  solution  colors 
cytisine  and  its  salts  blood  red.  Dilution  with  water,  or  acidi- 
fication, discharges  this  color.  Hydrogen  peroxide  also  pro- 
duces the  same  result.  The  solution  containing  hydrogen 
peroxide,  warmed  upon  the  water-bath,  becomes  intensely 
blue. 

2.  A.  Ramverda's^  Test. — A  Httle  nitrobenzene,  containing 
dinitro-thiophene,  poured  upon  cytisine  gives  a  fairly  stable, 
brilliant  red-violet  color. 

A  similar  color  given  by  coniine  is  very  unstable. 

3.  Nitro-Nitroso-Cjrtisine  Test. — Nitro-nitroso-cytisine  (see 
above),  formed  by  concentrated  nitric  acid,  serves  to  detect 
small  quantities  of  this  alkaloid.  Nitro-nitroso-cytisine  dis- 
solves with  difficulty  in  94  per  cent,  alcohol  and  crystallizes 
from  this  solvent  in  microscopic  prisms.  Flat,  tabular  crystals 
form  from  50  per  cent,  alcohol  which  is  a  better  solvent.  The 
solubility  of  nitro-nitroso-cytisine  in  concentrated  hydrochloric 
acid  indicates  basic  properties,  but  they  are  feeble,  for  dilution 
with  water  precipitates  this  compound  unchanged. 

^  Berichte  der  Deutschen  pharmazeutischen  Gesellschaft,  5,  267  (1895). 
^  Chemisches  Zentral-Blatt,  1900,  II,  268. 


200  DETECTION   OF  POISONS 

THE  DIGITALIS  GLUCOSIDES 

The  digitalis  plant  (Digitalis  purpurea  L.)  contains  in  all  its 
parts,  but  especially  in  the  leaves  and  seeds,  medicinally  useful 
substances  belonging  to  the  glucoside  group.  Thus  far  three 
digitalis  glucosides  have  been  isolated  as  well  characterized, 
crystalline  compounds  of  homogeneous  composition.  These  are 
digitahn  in  a  narrower  sense  (=  Digitahnum  verum  crystal- 
Hsatum  Kiliani)  C35H56O14;  digitoxin^  C34H54O11;  and  digitonin, 
C55H94O28  or  C54H92O28.  A  fourth  glucoside  called  digitalein 
seems  not  to  have  been  obtained  wholly  pure  as  yet. 

Digitonin,  C55H94O28  or  C54H92028,^  occurs  almost  exclusively 
in  digitalis  seeds,  the  leaves  containing  at  most  only  traces. 
Digitonin,  classified  at  present  with  the  saponins  (see  page  213), 
crystallizes  from  alcohol  in  fine  needles  soluble  in  50  parts  of 
50  per  cent,  alcohol.  Even  very  dilute  hydrochloric  acid 
hydrolyzes  digitonin  into  digitogenin,  dextrose  and  galactose  •} 

C66H94O28  +  2H2O  =  Csi.HsoOe  +  2C6H12O6  +  aCeHiaOeC?). 
Digitonin  Digitogenin        Dextrose  Galactose 

Digitonin  crystallizes  from  alcohol  in  fine  needles  which  soften 
at  235°  and  become  yellow.  Digitonin  is  not  a  cardiac  poison. 
Pure  digitonin  and  concentrated  sulphuric  acid,  upon  addition 
of  a  little  bromine  water,  give  a  color  which  becomes  intensely 
red. 

Digitoxin,  C34H54O11,  occurs  almost  exclusively  in  digitalis 
leaves.  This  very  active  and  highly  toxic  compound  is  almost 
wholly  insoluble  in  water  and  ether  but  soluble  in  alcohol  and 
chloroform.  Consequently  ether  precipitates  it  from  chloro- 
form solution.  Digitoxin  crystalhzes  from  85  per  cent,  alcohol 
in  leaflets  melting  at  145°.  Alcoholic  hydrochloric  acid  hy- 
drolyzes it  forming  digitoxigenin  and  digitoxose : 

C34H54O11  +  H2O  =  C22H32O4.+  2C6H12O4. 

Digitoxin  Digitoxigenin    Digitoxose 

Digitoxin  dissolves  in  concentrated  sulphuric  acid  with  a 
brownish  or  greenish  brown  color  unchanged  by  bromine. 

1  The  results  obtained  by  A.  Windhaus  (Berichte  der  Deutschen  chemischen 
Gesellschaft  42,  238  (1909)  favor  the  formxila  C66H94O28  for  digitonin. 

2  H.  Kiliani,  Berichte  der  Deutschen  chemischen  Gesellschaft  24,  340  (1891). 


POISONS  NOT  IN  THE  THREE  MAIN  GKOUPS      201 

Kiliani's  Digitoxin  Test.^ — Dissolve  a  trace  of  digi toxin  in 
3-4  cc.  of  glacial  acetic  acid  containing  iron  (loo  cc.  of  glacial 
acetic  acid  and  i  cc.  of  5  per  cent,  ferric  sulphate  solution). 
Cautiously  add  sulphuric  acid  containing  iron  (100  cc.  of  sul- 
phuric acid  and  i  cc.  of  5  per  cent,  ferric  sulphate  solution)  in 
about  the  same  quantity  as  an  under  layer.  A  dark  zone  ap- 
pears where  the  two  solutions  meet,  above  which  after  a  few 
minutes  a  blue  band  is  visible.  After  some  time  the  entire 
acetic  acid  layer  becomes  deep  indigo -blue. 

Digitalin,  C35H56O14,  according  to  Kiliani  occurs  only  in 
digitalis  seeds.  It  is  soluble  in  water  (i  :  1000)  and  very  active. 
Boiling  with  very  dilute  hydrochloric  acid  hydrolyzes  its  alco- 
holic solution  into  digitaligenin  and  two  sugars,  namely,  dex- 
trose and  digitalose:^ 

C36H50O14  +  H2O  =  C22H30O3  +  C6H12O6  +  2C7H14O6 

Digitalin  Digitaligenin  Dextrose        Digitalose 

Test  for  digitalin  as  follows: 

1.  Concentrated  sulphuric  acid  colors  pure  digitalin  orange 
yellow.  This  solution  soon  becomes  blood  red,  changing  upon 
addition  of  a  little  bromine  water  to  cherry  and  blue-red.  A 
drop  of  nitric  acid  or  ferric  chloride  solution  will  do  as  well  as 
bromine  water.  This  test  after  1-2  hours  is  surer  and  more 
permanent,  if  a  trace  of  digitalin  is  dissolved  direct  in  concen- 
trated sulphuric  acid  and  nothing  else  is  added. 

2.  Concentrated  hydrochloric  acid  dissolves  digitalin  mth  a 
golden  yellow  color,  changing  with  heat  to  garnet  or  violet-red. 

At  present  nothing  definite  is  known  regarding  the  fate  of  digitalis  glucosides 
in  the  human  organism,  or  the  products  into  which  they  are  changed  or  the 
forms  in  which  they  are  eliminated.  In  the  case  of  human  beings  elimination 
of  the  three  active  substances  has  never  been  observed.  Moreover,  R.  Kobert 
has  not  been  able  to  detect  anything  active  in  the  urine  of  animals  except  in 
isolated  cases.  Thus  far  it  has  not  been  possible  to  find  any  of  the  digitalis 
compounds  mentioned  above  in  blood  or  animal  organs.  In  a  toxicological 
analysis  especial  attention  would  have  to  be  given  to  vomitus  and  the  contents 
of  the  gastro-intestinal  tract.  But  there  is  slight  chance  of  detecting  the  digitalis 
bodies  in  such  material. 

^  Archiv  der  Pharmazie  234,  273-277  (1S96). 

2  H.  Kiliani,  Berichte  der  Deutschen  chemischen  GeseUschaf 1 3 1 ,  2454  (1S98). 


202  DETECTION   OE  POISONS 

ERGOT 

Officinal  ergot  (Secale  cornutum)  is  the  sclerotium  (compact  mycelium  oi 
spawn)  of  Claviceps  purpurea  collected  from  rye  shortly  before  the  fruiting  period 
and  dried  at  gentle  heat.  Ergot  is  commonly  known  as  an  abortifacient  and  in- 
toxications have  occurred  frotti  its  use.  Consequently  examinations  for  legal  pur- 
poses ma.y  require  its  detection  in  powders  and  other  mixtures.  Our  knowledge 
of  the  constituents  of  ergot  is  still  very  defective  notwithstanding  several  ex- 
haustive investigations.  Ergot  alkaloids,  as  ergotine,  ergotinine,  cornutine,  picro- 
sclerotine,  were  described  long  ago.  But,  with  the  possible  exception  of  ergotinine 
(Tanret,  C,  C.  Keller),  the  preparations  were  not  entirely  pure.  Ergot  contains 
in  addition  to  alkaloids  other  peculiar  chemical  substances  which  have  received 
but  little  attention.  They  have  not  the  characteristic  physiological  action  of 
ergot  but,  like  the  pigment  sclererythrin,  are  useful  for  purposes  of  identification. 
Among  these  substances  belong  sphacelic  acid  and  sclerotic  acid,  according  to  R. 
Kobert  a  very  poisonous  resin  having  acid  properties. 

Alkaloids. — The  most  recent^  researches  upon  ergot  mention  as  well  character- 
ized bases  ergotinine,  CasHsgNBOs,  and  hydro-ergotinine,  C3BH41N6O6.  Barger 
calls  the  latter  ergotoxine,  Ergotinine  crystalUzes  from  alcohol  in  long  needles 
melting  at  about  229°  when  heated  rapidly.  This  compound  dissolves  in  52 
parts  of  boiling  alcohol;  in  i.i  parts  of  ether;  and  is  readily  soluble  in  chloroform. 
Crystalline  salts  of  this  base  have  not  yet  been  prepared.  Hydro-ergotinine 
(  =  hydrate  of  ergotinine),  obtained  as  a  crystalline  phosphate  from  ergotinine 
mother  liquors  by  means  of  alcohol  and  phosphoric  acid,  is  a  white  powder  soften- 
ing at  155°  and  melting  at  162-164°.  Though  freely  soluble  in  alcohol,  it  dis- 
solves but  slightly  in  ether.  As  a  rule  the  salts  of  hydro-ergotinine  (ergotoxine) 
crystallize  well.^  By  preparing  a  cold  methyl  alcohol  solution  of  hydro-ergotinine 
and  boiling  this  solution  for  several  hours  under  a  return  condenser,  F.  Kraft  has 
converted  this  substance  completely  into  ergotinine.  On  the  other  hand,  ergoti- 
nine in  dilute  acetic  acid  solution  passes  back  almost  entirely  into  hydro-ergotinine 
within  10  days.  As  an  indication  of  purity,  a  solution  of  hydro-ergotinine  in  2 
parts  of  cold  methyl  alcohol  after  several  days  standing  should  not  deposit 
crystals  (ergotinine)  nor  become  green.  Solutions  of  both  alkaloids  are  fluores- 
cent. According  to  Keller  the  play  of  colors  with  sulphurc  acid  and  ferric  chloride 
is  characteristic  of  ergotinine  (see  below). 

Physiological  Action  of  the  Alkaloids.^ — Ergotinine  and  hydro-ergotinine  accord- 
ing to  A.  Jaquet  produce  convulsions  and  gangrene.  They  are  not,  however,  the 
cause  of  the  specific  uterine  contraction  characteristic  of  ergot.  Keller's  cornutine 
according  to  Kraft  is  identical  with  ergotinine,  according  to  G.  Barger  and  H.  H. 
Dale'  with  ergotimne,  which  is  impure  from  ergotoxine  (hydro-ergotinine). 
The  English  investigators  believe  that  the  physiological  effects  observed  with 
ergotinine  are  due  to  adhering  ergotoxine.  The  latter  is  readily  formed  when  the 
difficultly  soluble  ergotinine  is  brought  into  solution  by  means  of  glacial  acetic  acid, 
phosphoric  acid,  or  a  little  sodium  hydroxide  solution.     Ergotoxine  according  to 

^  F.  Kraft,  Archiv  der  Pharmazie  244,336  (1906)  and  G.  Barger,  Journal  of  the 
Chemical  Society  91,  337. 
^  G.  Barger  and  F.  H.  Carr,  Proceedings  of  the  Chemical  Society  23,  27 
^  Bio-Chemical  Journal  2,  240. 


POISONS   NOT   IN   THE    THREE    MAIN   GROUPS  203 

Barger  and  Dale  produces  the  effects  typical  of  ergot,  causing  powerful  contrac- 
tion of  the  uterus  and  later  abortion. 

Sclererythrin.' — This  is  the  pigment  of  the  outer  coat  of  ergot.  E.  Schniidt 
gives  the  following  directions  for  its  isolation.  Extract  freshly  powdered  ergot 
with  ether  to  remove  fat.  Then  moisten  the  powder  with  water  containing  tar- 
taric acid,  dry  and  extract  with  95  per  cent,  alcohol.  Filter  and  distil  the  alcohol. 
Extract  the  residue  with  ether.  This  solvent  now  dissolves  sclererythrin  which 
can  be  precipitated  by  means  of  petroleum  ether. 

Sclererythrin  is  an  amorphous  red  powder  which  can  be  sublimed.  It  is  in- 
soluble in  water  but  soluble  in  absolute  alcohol  and  glacial  acetic  acid.  This 
substance  behaves  like  an  acid,  dissolving  in  caustic  alkalies,  ammonia,  and 
alkaline  carbonate  and  bicarbonate  solutions  with  a  red  or  red-violet  color. 
Owing  to  presence  of  sclererythrin,  ether,  if  shaken  with  powdered  ergot  moistened 
with  tartaric  acid  solution,  becomes  red.  If  such  an  ether  solution  of  sclerery- 
thrin is  shaken  with  sodium  hydroxide  solution,  the  pigment  dissolves  in  the 
latter  which  then  becomes  red.  Solutions  of  this  pigment  show  characteristic 
absorption  bands  in  the  spectrum.  Moreover  the  pigment  gives  blue-violet 
precipitates  with  solutions  of  calcium  hydroxide,  barium  hydroxide  and  lead 
acetate.  The  precipitate  with  stannous  chloride  is  currant-red;  with  copper  sul- 
phate a  pure  violet;  with  ferric  chloride  a  deep  green;  and  with  chlorine  or  bromine 
water  a  lemon- yellow. 

Detection  of  Ergot  in  Flour,  Bread  and  Powders 

This  examination  usually  consists  in  undertaking  to  detect 
by  chemical  and  physical  means  the  red  pigment  sclererythrin 
which  is  characteristic  of  ergot.  The  property  possessed  by 
this  substance  of  passing  from  ether  into  a  solution  of  an  alkaline 
hydroxide  or  bicarbonate  is  especially  valuable  for  purposes  of 
identification, 

I.  Detection  of  Sclererythrin. — Shake  frequently  and  let 
10  grams  or  more  of  flour  stand  for  a  day  in  a  closed  flask  with 
20  cc.  of  ether  and  about  15  drops  of  dilute  sulphuric  acid  (i  :  5). 
Then  pass  the  ether  through  a  dry  paper,  wash  the  residue  with 
a  little  ether  and  shake  the  filtrate  thoroughly  with  ia-15  drops 
of  cold  saturated  sodium  bicarbonate  solution.  If  the  flour  con- 
tains ergot,  the  aqueous  layer  separates  with  a  violet  color. 

R.  Palm  extracts  the  flour  at  30-40°  with  10-15  times  its 
volume  of  40  per  cent,  alcohol  containing  a  few  drops  of  am- 
monia. Express  the  liquid,  filter  and  add  basic  lead  acetate 
solution  to  the  filtrate.  Press  the  precipitate  between  filter 
paper  and  warm  while  still  moist  with  a  little  cold  saturated 


204  DETECTION   OF  POISONS 

borax  solution.     A  red-violet  color  appears,  if  the  flour  con- 
tains ergot.    • 

2.  Spectroscopic  Examination. — This  test  gives  a  positive 
result,  if  the  material  (powdered  ergot,  flour,  bread)  contains 
more  than  o.i  per  cent,  of  ergot.  Examine  spectroscopically 
the  alkaline  and  acid  solution  of  the  pigment.  The  red  solu- 
tion, prepared  in  Test  i  by  means  of  ether  containing  sulphuric 
acid,  shows  two  absorption  bands.  One  lies  in  the  green  be- 
tween E  and  F  but  nearer  E  and  a  second  broader  band  in  the 
blue  midway  between  F  and  G.  Then  render  the  solution 
alkaline  with  ammonia.  Three  absorption  bands  should  ap- 
pear. The  first  lies  between  D  and  E,  the  second  at  E  some- 
what to  the  right  and  the  third  to  the  left  of  F. 

3.  Choline. — Ergot  powder,  warmed  with  dilute  potassium 
hydroxide  solution,  gives  the  characteristic  odor  of  trimeth- 
ylamine,  (CH3)3N,^  due  to  decomposition  of  choline  in  ergot. 

/CH2.CH2.OH 

(CH3)3N< 

\0H 

Occasionally  flour  that  does  not  contain  ergot  may  give  an 

odor  when  heated  with  potassium  hydroxide  solution. 

4.  Detection  and  Quantitative  Estimation  of  Ergotinine  (C.  C. 
Keller). — Dry  finely  powdered  ergot  over  lime,  place  25  grams 
in  a  Soxhlet  tube  and  completely  extract  fat  with  petroleum 
ether.  Dry  the  powder  at  gentle  heat,  add  100  grams  of  ether 
and  after  10  minutes  shake  well  with  milk  of  magnesia  (  i  gram 
of  MgO  and  20  cc.  of  water).  Shake  repeatedly  during  an  hour 
and  then  pass  80  grams  of  the  ether  solution  ( =  20  grams  of 
ergot)  through  a  covered  folded  filter  into  a  separating  funnel. 
Shake  the  ether  in  succession  with  25,  15,  10  and  5  cc.  of  0.5 
per  cent,  hydrochloric  acid.  Pour  the  hydrochloric  acid  ex- 
tracts, now  containing  the  ergot  alkaloids,  through  a  small 
moistened  filter,^     Add  ammonia  until  alkaline  and  extract 

^  The  so-called  corn  smut  (Ustilago  Maidis),  said  to  cause  effects  similar  to  those 
of  ergot,  also  gives  the  trimethylamine  odor  when  warmed  with  potassium  hydrox- 
ide solution,  for  it  contains  appreciable  quantities  of  choline. 

^  Clarify  the  filtrate  from  these  hydrochloric  acid  extracts,  if  not  clear,  by 
agitation  with  a  httle  talcum  powder,  previously  treated  with  hydrochloric  acid 
and  thoroughly  washed  with  water.     Then  filter  again.^ 


POISONS    NOT   IN    THE    TIIKEE   MAIN    GROUPS  205 

the  solution  twice  with  about  half  its  volume  of  ether.  J.et 
the  ether  extract  settle  in  a  dry  flask,  then  filter  into  a  dry 
weighed  flask  and  wash  the  filter  with  a  little  ether.  Distil 
the  ether  and  dry  flask  and  residue  at  ioo°  to  constant  weight. 
Good  German  ergot  contains  0.13-0.16  per  cent,  and  Russian 
ergot  0.22-0.25  per  cent,  of  the  alkaloid. 

To  detect  ergot  alkaloid  qualitatively,  proceed  as  follows: 

(a)  Dissolve  a  part  of  the  residue  in  i  cc.  of  concentrated 
sulphuric  acid  and  add  a  trace  of  ferric  chloride  solution.  The 
mixture  is  orange-red  and  becomes  at  once  deep  red  but  the 
margin  appears  bluish  to  bluish  green. 

(b)  Dissolve  a  part  of  the  residue  in  about  4  cc.  of  glacial 
acetic  acid  and  add  a  trace  of  ferric  chloride  solution.  Cau- 
tiously add  this  mixture  to  concentrated  sulphuric  acid  as  an 
upper  layer.  If  ergotinine  is  present,  a  brilliant  violet  color 
appears  where  the  two  liquids  meet. 

OPIUM 
Detection  of  Meconic  Acid  and  Meconine 

Since  it  is  comparatively  easy  to  procure  small  quantities  of 
opium  preparations,  especially  the  tincture,  poisoning  from  this 
source  is  possible.  Consequently,  it  is  often  desirable  to 
recognize  the  presence  of  opium  itself.  Detection  of  the 
alkaloids  narcotine  and  morphine,  always  present  in  opium  in 
considerable  quantity,  affords  partial  evidence  of  the  presence 
of  this  substance.  Moreover,  opium  always  contains  two  non- 
basic  substances,  meconic  acid  and  meconine.  Detection  of 
these -two  compounds  in  conjunction  with  narcotine  and 
morphine  definitely  determines  the  presence  of  opium. 

Meconic  Acid,  C7H4O7  =  C3H02(OH)(COOH)2,  is  an  oxy- 
pyrone-dicarboxylic  acid  (II)  and  therefore  a  derivative  of 
pyrone  (I) : 


0 

0 

c 

c 

/\ 

/\ 

HC      CH 

HC      C— OH 

I.       II       II 

11. 

II       II 

HC      CH 

HOOC— C      C— COOH 

\/ 

\/ 

0 

0 

Pyrone 

Meconic  acid 

206  DETECTION   OP   POISONS 

Meconic  acid  crystallizes  in  plates  or  prisms  with  3  mole- 
cules of  water.  It  is  easily  soluble  in  hot  water  and  alcohol. 
A  solution  of  a  ferric  salt  turns  a  meconic  acid  solution  dark  red. 

To  detect  meconic  acid,  extract  a  portion  of  the  material 
with  alcohol  containing  a  few  drops  of  hydrochloric  acid.  Filter 
and  evaporate  the  filtrate  upon  the  water-bath.  Dissolve  the 
residue  in  a  little  water  and  heat  the  filtered  solution  to  boil- 
ing with  excess  of  calcined  magnesium  oxide.  The  solution 
contains  magnesium  meconate.  Filter  hot  to  remove  undis- 
solved magnesium  oxide,  evaporate  the  filtrate  to  a  small 
volume  and  acidify  faintly  with  dilute  hydrochloric  acid.  Add 
a  few  drops  of  ferric  chloride  solution.  A  blood-red  color 
appears,  if  meconic  acid  is  present.  Warming  with  hydro- 
chloric acid  does  not  discharge  this  red  color,  in  which  respect 
it  differs  from  the  red  color  caused  by  acetic  acid.  This  color 
differs  from  that  caused  by  sulphocyanic  acid  in  not  being 
affected  upon  addition  of  gold  chloride.  But  stannous  chloride 
reduces  ferric  to  ferrous  oxide  and  discharges  the  color.  Nitrous 
acid,  however,  at  once  restores  it. 

These  tests  permit  the  identification  of  meconic  acid  in  an 
extract  from  only  0.05  gram  of  opium. 

Meconine,  C10H10O4. — Opium  contains  only  0.05-0.08  per 
cent,  of  this  compound.  It  forms  small  prisms,  melting  at  102° 
and  subliming  at  higher  temperature  without  decomposition. 
Meconine  dissolves  freely  in  alcohol,  ether,  benzene  and. chloro- 
form, but  less  easily  in  water.  Alkalies  convert  meconine 
into  easily  soluble  salts  of  meconinic  acid,  C10H12O5.  This 
monobasic  acid  cannot  exist  free  but  changes  to  meconine  when 
liberated  from  its  salts  by  a  mineral  acid.  Meconine,  formed 
by  abstracting  a  molecule  of  water  from  meconinic  acid,  is 
therefore  the  internal  anhydride  (lactone)  of  meconinic  acid: 
CHo— 6iH  CH2— O 


C 


c 


HC     C— CO  ;0H  HC     C— CO 


HC      C— OCH3  HC      C— OCH3 

\/  V 

C  C 

I  I 

0CH3  0CH3 

Meconinic  acid  Meconine 


POISONS    NOT   IN   THE    THREE    MAIN   GROUPS 


207 


To  detect  meconine,  extract  the  material  with  alcohol  con- 
taining sulphuric  acid.  Filter  and  evaporate  the  filtrate  to  a 
syrup  upon  the  water-bath.  Dissolve  the  residue  in  water  and 
extract  meconine  from  this  acid  solution  with  benzene.  Evapo- 
ration of  the  solvent  frequently  gives  crystals  of  meconine.  To 
detect  meconine,  dissolve  in  a  little  concentrated  sulphuric  acid. 
The  solution  is  green  but  turns  red  within  two  days.  If  the 
green  sulphuric  acid  solution,  or  that  which  has  turned  red  upon 
standing,  is  carefully  warmed,  a  fine  emerald-green  color  ap- 
pears, passing  through  blue,  violet  and  finally  back  to  red. 


Selenious-Sulphuric  Acid  Reagent  for  Opium  Alkaloids^ 

Prepare  the  reagent  used  in  these  tests  by  dissolving  0.5 
gram  of  selenious  acid  (H2Se03)  in  100  grams  of  pure  concen- 
trated sulphuric  acid.  This  reagent  is  especially  delicate,  for 
opium  alkaloids,  detecting  even  traces  of  morphine  and  codeine 
(0.05  milligram),  as  well  as  of  papaverine  (o.i  milligram). 
Selenious-sulphuric  acid  gives  the  following  color  reactions 
with  the  commoner  opium  alkaloids: 


Cold 

Hot 

Morphine 

Blue;  then  permanent  blue 
green  to  olive  green. 

Dark  blue  violet. 

Blue  quickly  changing  to 
emerald  green  and  later  to 
permanent  olive  green. 

Faint  greenish  yellow;  then 
violet. 

Greenish  steel  blue;  later 
cherry-red. 

Greenish,  dark  steel  blue; 
then  deep  violet. 

Deep  orange  graduallj^  fad- 
ing. 

Brown. 

Apomorphine    

Codeine 

Gradually  dark  brown. 
Steel  blue;  then  brown. 

Narceine 

Dark  violet. 

Narcotine 

Papaverine 

Thebaine 

Cherrj'  red. 
Intense  dark  violet. 
Dark  brown. 

^  Mecke,  Zeitschrift   fiir  offentliche  Chemie  5,  350  (1S99)  and  Zeitschrift  fiir 
analytische  Chemie  39,  468  (1900). 


H     H 

C      C 

/\/\ 

HsC.O- 

-C      C      CH 

1        II        1 

H3C.O- 

-C      C      N 

\/\^ 

c    c 

H      i 

CH2 

C 

/\ 

HC      CH 

1        II 

HC      C— 

\/ 

C 

208  DETECTION   OE   POISONS 


PAPAVERINE 

Papaverine,  C20H21NO4,  constitutes  about  0.5 — i  per  cent,  of  opium.  When 
crude  it  is  usually  mixed  with  narcotine.  To  remove  the  latter,  prepare  the  acid 
oxalate  of  papaverine  which  dissolves  with  diffi- 
culty in  water.  Crystallize  this  salt  from  boiling 
water  until  it  dissolves  in  concentrated  sulphuric 
acid  without  color.  Convert  papaverine  oxalate 
into  the  hydrochloride  by  treatment  with  calcium 
chloride  and  then  liberate  the  alkaloid  with  am- 
monia. This  product  crystallized  from  alcohol  is 
pure  papaverine. 

Papaverine  crystallizes  in  colorless  prisms  melt- 
ing at  147°.  This  alkaloid  is  insoluble  in  water; 
soluble  with  difficulty  in  ether  (1:260),  cold 
alcohol  and  benzene;  but  freely  soluble  in  hot 
-O  CH  ^-Icohol,  acetone  and  chloroform.  These  solutions 
are  neutral,  not  bitter,  and  optically  inactive. 
Papaverine  is  a  weak  base  which  dissolves  in  but 
I  does  not  neutralize  acetic  acid.      Ether  partially 

'  extracts  it  from  an  aqueous  tartaric  acid  solution 

and  completely  extracts  it  from  alkaline  solution.  Consequently  this  alkaloid 
appears  in  the  Stas-Otto  process  in  ether  extract  B.  Chloroform  extracts 
papaverine  with  almost  as  much  ease  from  an  acid  solution  as  from  one  that 
is  alkaline. 

Constitution. — Papaverine  is  a  monacid,  tertiary  base  which  combines  with 
alkyl  iodides  forming  crystalline  addition  products.  As  it  forms  no  acetyl  deriva- 
tive  with  acetic  anhydride,  free  hydroxyl  is  not  present.  But  there  are  probably 
four  methoxyl  groups,  for  it  loses  four  methyl  groups  when  treated  with  hydriodic 
acid  according  to  Zeisel's  method.  Consequently  all  the  oxygen  atoms  in  papa- 
verine are  present  as  methoxyl  groups.  The  researches  of  Guido  Goldschmiedt, 
extending  from  1883  to  1898,  have  completely  explained  the  constitution 
of  papaverine.  Moderate  oxidation  with  potassium  permanganate  and  sulphuric 
acid  gives  papaveraldine,  C20H19NO6,  without  breaking  the  carbon  chain.  Fusion 
with  potassium  hydroxide  breaks  the  latter  into  nitrogen-free  veratric  acid  and 
the  nitrogenous  base  dimethoxy-isoquinoline:^ 

1  Isoquinoline  (II)  is  isomeric  with  quinohne  (I)  and  Uke  the  latter  is  a  monacid, 
tertiary  base: 

I.  H     H  II.  H     H 

C      C  C      C 

HC      C      CH  HC      C      CH 


HC      C      CH 

HC      C      N 

\/\/- 

\/\/ 

'C     N 

c    c 

H 

H     H 

Quinoline 

Isoquinoline 

/ 

POISONS   NOT   IN   THE   THREE    MAIN   GROUPS  209 


H     H 

H     H 

C      C 

C      C 

/'\/\ 

/\/\ 

HaCO— C      C      CH 
HaCO— C      C      N 

liaCO— C      C      CH 

1       II       1 

1       II       1 
H3CO— C      C      N 

\/\/' 

VV 

C      C 

C      C 

H      1         H 

H     H 

C0+  OK 

Dimethoxy-isoquinol  ine 

1 

COOK 

C 

1 

/X 

c 

HC      CH 

/■x 

1        II 

HC      CH 

HC      C— OCH3 

1        II 

\/ 

HC      C— OCH3 

c 

\/ 

1 

C 

0CH3 

1 

OCH3 

Papaveraldin 

Veratric  acid 

Detection  of  Papaverine 

The  following  general  reagents  precipitate  papaverine  in  a 
dilution  of  i  :  10,000:  phospho-molybdic  acid,  potassium  bis- 
muthous  iodide  and  iodo-potassium  iodide. 

The  following  still  give  precipitates  in  a  dilution  of  i  :  5000: 
tannic  acid,  gold  chloride  and  potassium  mercuric  iodide. 

The  following  special  tests  should  be  made: 

1.  Concentrated  Sulphuric  Acid. — The  cold  colorless  solution 
of  papaverine  in  this  acid  becomes  dark  violet  upon  gentle 
warming.  But  even  a  cold  solution  of  impure  papaverine  in 
this  acid  is  violet. 

2.  Froehde's  Test. — The  solution  of  pure  papaverine  in  this 
reagent  is  green.  Blue,  violet  and  finally  a  briUiant  cherry  red 
color  appear  upon  warming  the  solution. 

3.  Solutions  of  this  alkaloid  in  concentrated  nitric  acid,  or 
Erdmann's  reagent,  are  dark  red. 

Heat  to  boiling  a  solution  of  i  part  of  papaverine  with  10  parts  of  nitric  add 
(sp.  gr.  1.06  =  10  per  cent.  HNO3).  As  the  solution  cools,  yellow  crystals  of 
the  nitrate  of  nitro-papaverine,  C2oH2o(N02)N04.HN03.H20,  appear.  Yellow 
prisms  of  nitro-papaverine,  C2oH2o(N02)X04.H:0,  may  be  obtained  from  this 
nitrate  by  means  of  ammonia. 

4.  Ammonia  colors  the  greenish  solution  of  papaverine  in 

14 


210  DETECTION   OF  POISONS 

chlorine   water  deep  red-brown  which  becomes  later  almost 
black-brown, 

5.  Selenious-Sulphuric  Acid  Test. — See  page  207  for  the 
color  changes  given  by  pure  papaverine  dissolved  in  this 
reagent. 


PILOCARPINE 

Pilocarpine,  C11H16N2O2,  occurs  with  isopilocarpine  and  probably  also  with 

pilocarpidine  in  the  leaves  of  jaborandum   (Pilocarpus  pennatifolius^).     The 

H       O  free  base  as  usually  obtained  is  semi-liquid,  viscous,  non- 

C2H6 — C — C  volatile  and  alkaline.      It  dissolves  but  slightly  in  water; 

\0       is  freely  soluble  in  alcohol,   ether   and   chloroform;   and 

jjQ Q  insoluble  in  benzene.     Solutions  of  pilocarpine  and  its  salts 

I  H2  are  dextro-rotatory.     This  alkaloid  is  a  strong  base  neu- 
CH2  tralizing   acids   and   forming   salts  that  are  usually  crys- 

/    ^  talline.     Caustic  alkalies,  added  to  concentrated  solutions 

Q jsj  of  pilocarpine  salts,  precipitate  the  free  base  which  redis- 

II  Xpi-cT  solves    in    an    excess  of    the    precipitant.       Solutions   of 
11        ^  sodium  hydroxide,  or  sodium  ethylate  (C2H6.0Na),  cause 

a  molecular  rearrangement  of  pilocarpine.  This  reaction 
runs  more  smoothly,  if  pilocarpine  hydrochloride  is  heated  for  half  an  hour  at 
200°.  The  product  of  this  change  is  isopilocarpine,  C11H16N2O2,  isomeric 
and  very  likely  stereo-isomeric  with  pilocarpine.  Both  isomeric  pilocarpines 
differ  in  melting  points,  solubilities  and  particularly  in  specific  rotation. 
Isopilocarpine  is  less  dextro-rotatory  than  pilocarpine  and  crystalUzes  in 
deliquescent  prisms  easily  soluble  in  water  and  alcohol.  The  salts  of  the 
two  bases  also  show  similar  differences: 

Pilocarpine  nitrate,       C11H16N2O2.HNO3;  mpt.  178°;  [a]D  =  +  82  .90°. 
Isopilocarpine  nitrate,  CHH16N2O2.HNO3;  mpt.  159°;  [q;]d  =  +  35  .68°. 

Jowett^  has  succeeded  in  converting  isopilocarpine  into  pilocarpine  by  means 
of  the  same  reagent  used  in  converting  pilocarpine  into  isopilocarpine.  Pure 
isopilocarpine,  heated  with  pure  alcoholic  potassium  hydroxide,  gives  a  mixture 
of  unaltered  isopilocarpine  and  pilocarpine.  The  identity  of  the  latter  with 
pure  pilocarpine  was  established  by  preparing  the  hydrochloride  and  nitrate 
(mpt.  178°).  This  reciprocal  conversion  of  one  alkaloid  into  the  other  strongly 
supports  the  idea  of  the  stereo-isomerism  of  pilocarpine  and  isopilocarpine. 
Pinner  was  the  first  to  show  that  the  two  nitrogen  atoms  of  the  two  isomeric 

^  According  to  Jowett,  jaborine,  which  has  been  described  as  another  alkaloid 
peculiar  to  jaborandum  leaves,  is  a  mixture  of  isopilocarpine,  pilocarpidine,  a 
little  pilocarpine  and  pigment. 

^  Proceedings  of  the  Chemical  Society  21,  172  (1905). 


POISONS  NOT  IN  THE  THREE  MAIN  GROUPS      211 

bases  belong  to  a  glyoxalinc  ring.^     In  1905  Jowctt  proposed  for  pilocarpine 
and  isopilocarpine  the  following  formula;: 

+       +  -        + 

C2H6.CH— CH— CH2— C— N— CI-I3     Calls.CH— CH— CII2— C— N— CH3 

I      I  11      /  I      I  II      / 

OC        CH2  liC— N  OC        CIi2  HC— N 


o  o 

Pilocarpine  Isopilocarpine 


Detection  of  Pilocarpine 

Ether,  chloroform  or  benzene  extracts  pilocarpine  from 
aqueous  solutions  alkaline  with  sodium  hydroxide  or  carbonate. 
Evaporation  of  these  solutions  leaves  a  thick,  non-crystalline, 
alkaline  syrup.  The  general  reagents  especially  delicate  for 
pilocarpine  are:  iodo-potassium  iodide,  phospho-molybdic  acid, 
phospho-tungstic  acid  and  potassium  bismuthous  iodide. 

I.  Place  a  particle  of  potassium  dichromate  and  1-2  cc.  of 
chloroform  in  a  test-tube.  Then  add  pilocarpine  itself,  or  its 
solution  and  i  cc.  of  3  per  cent,  hydrogen  peroxide.  Shake  for 
several  minutes.  The  mixture  yellowish  at  first  gradually 
darkens  and  in  5  minutes  is  dark  brown.  Depending  upon  the 
amount  of  pilocarpine,  the  chloroform  is  blue-violet,  dark  or 
indigo-blue.  But  the  upper  aqueous  solution  gradually  fades. 
The  chloroform  mixture  is  an  intense  blue  from  0.0 1  gram  of 
pilocarpine  and  blue-violet  from  o.ooi  gram  and  less.  The 
color  lasts  from  an  hour  to  a  day  (H.  Helch^). 

Apomorphine  (0.0 1  gram)  colors  chloroform  blue- violet  even  mthout  hydrogen 
peroxide.  Strychnine  gives  a  barely  perceptible  bluish  tint  which  changes  com- 
pletely within  a  few  minutes.  There  is  a  color  with  antipyrine  only  after 
acidification  of  the  hydrogen  peroxide. 

^  Glyoxaline,  or  imidazole  (C3H4N2),  is  obtained  by  the  action  of  ammonia 
upon  glyoxal  in  presence  of  formaldehj'de.     It  is  a  strong  base  and  cr\*stalluie. 

HC  =  I6 HsInh         H  HC— nh 

I  +  -t-     C        =         I        ^CH     +     3H2O. 

HC  =  \0ZZI^M}^^^^^^^^'^  HC— N 

Glyoxal  Formaldehyde  Glyoxaline 

^  Pharmazeutische  Post  35,  289,  498  (1902)  and  39,  313  (1906). 


212  DETECTION   OF   POISONS 

2.  Mandelin's  reagent  dissolves  pilocarpine  with  a  golden 
yellow  color  which  gradually  changes  to  bright  green  and  finally 
to  hght  brown. 

3.  The  solution  of  pilocarpine  in  formalin-sulphuric  acid 
becomes  yellow,  yellowish  brown  and  blood  red,  if  warmed. 

Thus  far  fatal  poisonings  from  this  alkaloid  have  not  oc- 
curred and  nothing  is  known  as  to  the  possibility  of  its  detection 
in  the  cadaver. 

PTOMAINES 

Ptomaines  are  basic  substances  containing  nitrogen  and  may  be  toxic  or 
non-toxic.  They  are  produced  during  putrefaction  of  cadavers  under  the  in- 
fluence of  bacteria.  They  are  to  be  regarded  to  some  extent  as  products  of 
bacterial  metabolism  and  are  nearly  always  present  in  cadavers,  especially  in 
those  parts  which  are  in  an  advanced  state  of  putrefaction.  Many  ptomaines 
closely  resemble  alkaloids.  Like  alkaloids  they  give  precipitates  with  the  general 
reagents,  and  certain  ptomaines  resemble  well-defined  alkaloids  even  with  special 
reagents.  Hence  ptomaines  are  of  very  great  importance  in  forensic  chemistry, 
since  their  presence  may  easily  lead  to  mistakes  and  false  conclusions.  These 
putrefactive  products  also  resemble  vegetable  bases  in  their  behavior  with  sol- 
vents. Ether  extracts  some  of  them  from  acid  solution  and  others  from  alkaline 
solution,  whereas  certain  ptomaines  are  removed  from  alkaline  solution  only  by 
amyl  alcohol  or  chloroform.  Most  of  the  ptomaines  are  strong  reducing  agents, 
for  example,  they  will  immediately  convert  potassium  ferricyanide  into  ferro- 
cyanide.  Consequently,  they  give  the  Prussian  blue  test  with  a  dilute  mixture 
of  solutions  of  ferric  chloride  and  potassium  ferricyanide.  Many  of  the  alkaloids 
like  morphine  resemble  the  ptomaines  in  this  respect. 

The  resemblance  of  a  ptomaine  to  a  definite  vegetable  base  is  frequently  con- 
fined to  some  one  reaction,  and  never  extends  to  all  the  reactions  characteristic 
of  the  particular  alkaloid.  In  a  legal-chemical  investigation  no  precaution, 
which  guards  against  mistaking  a  ptomaine  for  a  vegetable  base,  should  be 
omitted.  It  is  an  invariable  rule  to  make  every  test  characteristic  of  the 
suspected  alkaloid,  and  not  to  be  satisfied  with  possibly  one  positive  test.  A 
determination  of  the  physiological  action  of  the  substance  should  supplement 
the  chemical  examination.  A  ptomaine  may  resemble  a  vegetable  base  chemi- 
cally, and  yet  the  two  substances  may  differ  very  decidedly  in  physiological  action. 
Thus  far,  ptomaines  have  been  found  which  show  certain  resemblances  to  coniine, 
nicotine,  strychnine,  codeine,  veratrine,  delphinine,  atropine,  hyoscyamine, 
morphine  and  narceine.  Selmi  has  described  a  putrefactive  product  which 
resembles  morphine.  Ether  failed  to  extract  it,  either  from  acid  or  alkaline 
solution,  whereas  amyl  alcohol  removed  it  with  ease  from  an  alkaline  or  am- 
moniacal  solution.  It  liberated  iodine  from  iodic  acid,  but  failed  to  give  the 
tests  which  are  characteristic  of  morphine  alone,  namely,  Husemann's,  Pellagri's 
and  the  ferric  chloride  tests! 


POISONS   NOT   IN   Till-:    THREE    MAIN    GROUPS  2 1  'A 

The  object  in  such  cases  must,  be  to  get  a  result  about  which  there  can  be  no 
doubt.  Every  possible, means  must  be  used  to  isolate  the  alkaloid  in  a  perfectly 
pure  state.  When  this  can  be  accomplished,  the  nature  of  the  poison  can  always 
be  established  beyond  question. 

SAPONINS 

The  term  saponins,  or  saponin  substances,  includes  a  large  number  of  gluco- 
side-like  bodies  of  widespread  occurrence  in  the  vegetable  kingdom  and  having 
in  common  certain  chemical,  physical  and  especially  physicological  properties. 
Their  aqueous  solutions  when  shaken  foam  readily.  In  this  respect  they 
resemble  the  soaps.  Many  saponin  substances  have  a  sharp,  harsh  taste.  In 
powdered  form  they  excite  violent  sneezing.  They  are  capable  of  holding  many 
finely  divided  substances  in  a  state  of  emulsion.  They  dialj'ze  incompletely 
and  salts  precipitate  them  from  solution.  Excepting  the  gluco-alkalcid  sclanine, 
which  contains  nitrogen  and  is  alkaline,  the  saponins  may  be  classified  chemically 
as  nitrogen-free  glucosides.  Most  saponins  are  neutral  and  only  a  few  are 
faintly  acid.  Neutral  saponins  and  alkali  salts  of  acid  saponin  substances  dis- 
solve in  water  and  hot  aqueous  alcohol  but  are  insoluble  in  absolute  alcohol  and 
ether.  Barium  hydroxide  and  lead  acetate  (neutral  and  basic)  precipitate 
saponins  from  concentrated  aqueous  solution.  The  former  gives  baryta  saponins. 
Basic  lead  acetate  precipitates  all  saponins  but  the  neutral  salt  precipitates 
only  acid  saponins.  Ammonium  sulphate  is  capable  of  salting  saponin  sub- 
stances from  solution  as  it  does  proteins.  Solutions  of  saponins  in  concentrated 
sulphuric  acid  are  yellow,  gradually  becoming  red  and  sometimes  violet  and 
blue-green.  The  detection  thus  far  of  saponin  substances  in  more  than  50  plant 
families  having  over  200  monocotyledenous  and  dicotyledenous  species  shows 
the  wide  occurrence  of  these  substances  in  the  vegetable  kingdom.  Saponins 
occur  in  roots  (Senega,  Saponaria),  tubers  (Cyclamen),  barks  (Quillaja,  Guaia- 
cum),  fruits  (Sapindus,  Saponaria),  seeds  (^sculus,  Agrostemma,  Thea),  stems 
(Dulcamara)  and  leaves  (Guaiacum).  In  fact  almost  any  part  of  the  plant 
organism  may  contain  saponins.  The  plant  families,  producing  saponin  sub- 
stances in  greater  abundance,  are  the  sapindaceas,  caryophyllaceas,  colchicacese 
polygalaceee,  silenese  and  solanacese.  Quite  considerable  quantities  of  saponins 
may  occur  in  the  particular  part  of  the  plant.  , 

Saponin  solutions,  heated  with  dilute  hydrochloric  or  sulphuric  acid,  are 
hydrolyzed  into  sugars  and  a  non-toxic  substance  insoluble  in  water  called 
sapogenin.  The  sapogenins  have  not  been  extensively  investigated  but  they 
are  not  entirely  identical. 

The  following  saponins  have  been  more  closely  studied: 

Digitonin:  in  the  seeds  of  Digitalis  purpurea. 

Saponin:  in  the  root  of  Saponaria  officinalis  (4-5  per  cent.). 

Githagin:  in  the  seeds  of  the  corn  cockle,  Agrostemma  githago  (6.5  per 
cent.). 

Senegin:  in  Senega  root,  the  root  of  Polygala  senega. 

Struthiin:  in  levantine  soap  root,  the  root  of  Gj^sophila  struthium  (14  per 
cent). 

Quillaja-Sapotoxin:  in  the  bark  of  Quillaja  saponaria  (S.S  per  cent.). 


214  DETECTION   OF   POISONS 

Sapindus-Sapotoxin:  in  the  fruit  of  Sapindus  saponaria. 

Sarsaparilla-Saponin:  in  the  sarsaparilla  root,  the  root  of  various  kinds  of 
sniilax. 

Physiological  Action  of  Saponins. — Almost  without  exception  saponin  sub- 
stances are  highly  toxic,  if  introduced  directJy  into  the  blood.  Most  saponins 
are  absorbed  with  difficulty.  Consequently  healthy  individuals  may  take  dilute 
saponin  solutions  by  the  mouth  in  considerable  quantities  without  iU  effects. 
Toxic  saponins  act  in  common  as  protoplasmic  irritants.  In  larger  doses  saponin 
substances  kill  protoplasm.  They  manifest  in  various  ways  their  power  of  acting 
as  protoplasmic  poisons.  Saponins  act  upon  blood- corpuscles  for  the  same 
reason.  R.  Kobert  and  his  collaborators  have  shown  defibrinated  blood,  diluted 
loo  times  with  physiological  salt  solution  (see  below),  to  be  the  best  and  most 
convenient  reagent  for  saponin  substances.  Saponins  cause  haemolysis  and  the 
blood  solution  becomes  laky.  Agglutination  and  formation  of  methaemoglobin 
do  not  occur.  The  freer  the  blood  is  of  serum,  the  more  pronounced  the  hae- 
molytic  action  of  saponin  substances  upon  blood-corpuscles.  Recent  investi- 
gations have  shown  that  saponins  act  more  vigorously  upon  blood- corpuscles 
isolated  from  serum,  because  blood  serum  contains  cholesterin  which  has  a  pro- 
tective influence  and  retards  haemolysis.  Most  likely  the  haemolytic  action  of 
saponins  is  due  to  removal  of  cell  membrane  lecithin,  the  chief  constituent  of  the 
cell  wall,  from  red  blood-corpwscles,  for  lecithin-saponins  are  fortaed.  Saponins 
also  combine  with  cholesterin,  as  well  as  with  lecithin,  forming  cholesterin- 
saponins.  The  affinities  of  a  saponin  having  been  satisfied  by  cholesterin,  it  no 
longer  acts  upon  the  lecithin  of  the  membrane  of  blood- corpuscles.  Thus 
cholesterin  prevents  haemolysis,  which  a  saponin  may  produce,  and  so  acts  as 
an  antidote  to  saponin  substances.  Ransam^  has  made  the  important  discovery 
that  addition  of  cholesterm  checks  the  solvent  action  of  a  saponin  upon  blood- 
corpuscles.  At  first  it  was  not  known  whether  this  antidotal  action  was  due  to  a 
chemical  reaction,  or  to  adsorption,  that  is  to  say,  to  a  physical  process.  R. 
Kobert^  as  well  as  Madsen  and  Noguchi^  were  able  to  dissolve  cholesterin, 
which  is  insoluble  in  water,  in  an  aqueous  saponin  solution.  They  assumed  that 
this  physiologically  inactive  solution  contained  a  labile  saponin-cholesterin 
compound  no  longer  having  hemolytic  power.  Recently  A.  Windhaus*  has 
definitely  proved  that  saponin-cholesterides  exist.  Digitonin-cholesteride, 
C60H94O28.C27H46O,  crystallizes  in  fine  needles,  when  a  hot  alcoholic  solution  of 
digitonin  (i  molecule)  is  poured  into  a  similar  solution  of  cholesterin  (i  mole- 
cule). This  cholesteride  is  formed  without  elimination  of  water.  Hence  in  this 
reaction  between  digitonin  and  cholesterin  we  are  dealing  most  probably  with  the 
formation  of  a  molecular  compound. 

Saponin  solutions  also  dissolve  white  blood- corpuscles  but  only  at  higher 
concentrations.  A  physiological  action  characteristic  of  many  saponins  is 
exhibited  in  the  stupefaction  and  killing  of  fish,  even  in  water  containing  only 
1:200,000  of  saponin  substance  (R.  Kobert). 

^  Deutsche  medizinische  Wochenschrift  1901,  194. 

^  R.  Kobert,  Die  Saponine,  Stuttgart,  1904. 

^  Chemisches  Zentralblatt,  1905  I,  1265. 

^  Berichte  der  Dcutschen  chemischen  Gesellschaft  42,  238  (1909). 


POISONS  NOT  IN  THE  THREE  MAIN  GROUPS      215 

Detection  of  Saponins 

The  matter  of  solubility  is  especially  important  in  isolating 
saponin  substances  from  mixtures.  All  saponins  are  soluble  in 
water  and  some  in  alcohol,  but  they  are  practically  insoluble 
in  ether,  benzene,  chloroform  and  petroleum  ether.  Employ 
neutral  or  basic  lead  acetate  (see  above)  in  isolating  saponins. 
Decompose  the  washed  precipitate  with  hydrogen  sulphide, 
filter  and  evaporate  the  filtrate  upon  the  water-bath.  Pre- 
cipitate the  saponin  with  absolute  alcohol  and  ether  from  the 
concentrated  solution.  Solutions  of  most  saponins  in  con- 
centrated sulphuric  acid  are  red  or  yellowish  red,  gradually 
becoming  violet.  Saponin  substances  give  various  colors  with 
Froehde's  reagent  and  vanadic-sulphuric  acid:  brown,  red- 
brown,  blue,  green  and  violet  (see  solanin) .  A  saponin  solution , 
heated  with  dilute  hydrochloric  acid,  undergoes  hydrolysis  and 
then,  owing  to  formation  of  sugar,  reduces  Fehling's  solution 
with  heat. 

Detection  in  Foaming  Beverages  (Beer,  Wine,  Effervescing  Lemonade)  ^ 

Treat  the  beverage  to  be  tested  for  saponin  with  excess  of 
basic  magnesium  carbonate,  evaporate  to  about  loo  cc.  and  mix 
with  2  volumes  of  96  per  cent,  alcohol.  Filter  after  30  minutes 
and  evaporate  the  alcohol  from  the  filtrate.  Filter  the  residue 
hot  and  extract  the  cold  filtrate  with  sufficient  Hquid  carbolic 
acid^  to  leave  about  5  cc.  undissolved.  Add  ammonium  sulphate 
to  hasten  the  separation  of  the  carbolic  acid  layer.  Then  shake 
the  latter  with  water  and  a  mixture  of  2  volumes  of  ether  and  i 
volume  of  petroleum  ether.  Evaporate  the  aqueous  solution 
to  dryness  upon  the  water-bath.  Wash  the  residue  with  cold 
absolute  alcohol,  in  case  of  wine,  and  with  acetone,  in  case  of 
beer.  The  residue  fails  to  give  the  saponin  reaction  well, 
that  is  to  say,  a  red  color  with  concentrated  sulphuric  acid,  un- 
less treated  as  described.     E.  Schaer  dissolves  the  residue  in  con- 

^  K.  Brunner,  Zeitschrift  fiir  Untersuchung  der  Nahrungs-  und  Genussmittel 
S,  1197  (1902). 
^  Acidum  carboKcum  liquefactum  of  the  German  Pharmacopoeia. 


216  DETECTION   OF   POISONS 

centrated  aqueous  chloral  hydrate  solution  and  adds  the  latter 
to  concentrated  sulphuric  acid  as  an  upper  layer.  A  saponin 
produces  a  yellow,  then  purple-red  and  finally  mallow-blue 
zone. 

Detection  of  Githagin  (Com  Cockle  Saponin)  in  Flour 

Heat  500  grams  of  flour  with  i  liter  of  alcohol  (sp.  gr.  0.8496 
=  85  per  cent,  by  volume).  Filter  hot,  distil  most  of  the 
alcohol,  add  absolute  alcohol  as  well  as  ether  to  the  residue  and 
let  stand  12-24  hours.  Collect  the  precipitate  upon  a  filter  and 
dry  for  a  short  time  at  100°  to  coagulate  possible  protein. 
Dissolve  in  a  little  cold  water,  filter  and  precipitate  githagin 
from  the  filtrate  with  absolute  alcohol,  best  with  addition  of 
ether.  Githagin  thus  obtained  is  a  yellowish  white  powder 
having  a  sharp,  harsh  taste. 

To  prove  the  presence  of  a  saponin  substance,  agitate  its 
aqueous  solution  which  should  foam.  Then  heat  the  solution 
with  dilute  hydrochloric  acid  and  test  its  reducing  power  with 
FehUng's  solution.  Finally,  if  possible,  perform  the  physio- 
logical test  with  blood.  Dilute  defibrinated  ox  blood  with  100 
volumes  of  0.9  per  cent,  sodium  chloride  solution  and  add 
the  solution  of  supposed  githagin  in  0.9  per  cent,  sodium  chloride 
solution.  The  blood  solution  at  once  becomes  laky,  if  githagin 
is  present.  According  to  J.  Brandl,^  Agrostemma-Sapo toxin 
(githagin)  produces  haemolysis  in  very  great  dilution  (i :  50,000). 
But  after  previous  treatment  with  cholesterin,  even  o.oi  gram 
shows  no  haemolytic  action  whatever. 

Physiological  Salt  Solution  and  Haemolysis 

To  prevent  red  blood-corpuscles  from  changing  volume  in 
experiments  requiring  dilution  of  blood,  an  isotonic  salt 
solution  must  be  used.  What  is  an  isotonic  solution?  If  n 
gram-molecules  of  a  body  A  are  dissolved  in  a  definite  volume  of 
solvent  and  n  gram-molecules  of  a  body  B  are  dissolved  in  an 

^  Archiv  fiir  experimentelle  Pathologie  und  Pharmakologie,  54,  245. 


POISONS  NOT  IN  THE  THREE  MAIN  GROUPS      217 

equal  volume  of  the  same  solvent,  certain  properties  of  the 
original  solvent  are  changed  equally  in  both  cases.  The  freezing 
point  of  the  solutions  is  lowered  and  the  boiling  point  raised 
equally.  The  two  solutions  have  the  same  vapor  tension  and 
the  same  osmotic  pressure.  In  other  words  they  are  isotonic. 
Blood-corpuscles  retain  their  volume  unchanged,  if  brought 
into  a  salt  solution  having  the  same  osmotic  pressure  as  the 
blood  serum.  Such  a  salt  solution  is  isotonic  with  blood  serum. 
In  the  case  of  human  and  mammalian  blood  an  isotonic  solution 
of  sodium  chloride  has  a  concentration  of  9  per  thousand  = 
physiological  salt  solution.  Such  a  solution  formerly  contained 
0.6  per  cent,  of  sodium  chloride.  Blood-corpuscles  give  up 
water  to  solutions  of  higher  concentration  than  0.9  per  cent. 
NaCl  (hyperisotonic  solutions)  until  osmotic  equilibrium  is 
established.  They  shrivel  and  hence  have  a  smaller  volume. 
On  the  other  hand,  blood-corpuscles  in  salt  solutions  of  lower 
concentration  (hypisotonic  solutions)  take  up  water  and  become 
distended.  In  diluting  blood  with  water,  this  swelling  may 
go  far  enough  to  cause  hsemoglobin  to  separate  from  the  stroma 
and  pass  into  the  aqueous  solution.  This  process  is  called 
haemolysis..  Alternate  freezing  and  thawing  of  blood  may 
produce  haemolysis.  Various  chemical  substances,  which  act  as 
protoplasmic  poisons,  cause  the  same  result.  Such  substances 
are  ether,  alcohol,  chloroform,  alkalies,  gallic  acids,  solanine, 
etc.  The  saponins  described  above  are  also  powerful  hsemolytic 
agents.  Finally,  those  globulicidal  substances,  or  haemolysins, 
normally  occurring  in  blood  sera,  as  well  as  those  produced  in 
immunization,  belong  in  this  class. 

SOLANINE 

Solanine,  C62H93NO1S,  at  the  same  time  an  alkaloid  and  a  glucoside  (gluco- 
alkaloid)  occurs  in  the  potato  plant  (Solanum  tuberosum)  and  in  other  Solanaceae 
as  Solanum  nigrum,  Solanum  dulcamara  and  Solanum  Ijxopersicum  (tomato). 
It  has  been  found  also  in  Scopoliaceae,  as  in  Scopolia  orientaJis  and  Scopolia 
atropoides.  Solanine  is  not  uniformly  distributed  in  all  parts  of  the  potato  plant 
but  is  most  abundant  in  the  berr3'-Hke  fruit  and  m  the  chlorophj'll-free  sprouts 
appearmg  m  the  spring  upon  potatoes  that  he  ia  a  cellar.  Schmiedeberg  and 
Meyer  found  0.024  gram  of  solanine  per  kilogram  of  peeled  potatoes  in  Januarj-  and 


218  DETECTION   OP   POISONS 

February  but  0.044  gram  in  unpeeled  potatoes.  Potato  peelings  gave  0.71  gram 
of  solanine  per  kilogram  and  potato  sprouts  i  cm.  long  even  5.0  grams.  The 
appearance  of  solanine  according  to  R.  Werk  is  due  to  the  life  processes  of  Bac- 
terium solaniferum  (?). 

Solanine  crystallizes  in  white  needles  having  a  bitter  taste  and  melting  at 
244°.  Even  boiling  water  dissolves  only  a  little  of  this  alkaloid  (about  i:  8000). 
It  is  soluble  in  500  parts  of  cold  and  125  parts  of  boiling  alcohol;  and  in  about 
4000  parts  of  ether.  These  solutions  are  faintly  alkaline.  Hot  saturated  solutions 
of  solanine  in  alcohol  and  amyl  alcohol  gelatinize  upon  cooling.  Ether,  chloro- 
form and  benzene  do  not  extract  solanine  either  from  acid  or  alkaline  solution. 
But  hot  amyl  alcohol  extracts  solanine  from  acid  solution  and  from  solutions  al- 
kaline with  sodium  hydroxide  or  ammonia.  Solanine  is  a  weak  base,  readily  dis- 
solving in  acids,  as  acetic  acid,  and  forming  crystalline  salts.  Dilute  hydrochloric 
or  sulphuric  acid  hydrolyzes  solanine  to  solanidine,  C40H61NO2,  galactose  and 
rhamnose.  Hydrolysis  is  very  slow  in  the  cold  but  rapid  upon  heating.  The 
hydrochloride  or  sulphate  of  solanidine  separates  as  a  difficultly  soluble,  crys- 
talline powder.  A  good  yield  of  solanidine  is  obtained,  according  to  Wittmann, 
by  heating  solanine  under  a  return-condenser  with  10  times  the  quantity  of  2 
per  cent,  sulphuric  acid,  until  the  liquid  is  yellowish  and  the  filtrate  upon  further 
boiling  no  longer  deposits  solanidine  sulphate.  Solanidine,  precipitated  from  its 
sulphate  with  ammonia  and  recrystallized  from  ether,  forms  colorless,  silky 
needles,  melting  at  207°  and  dissolving  with  difficulty  in  water  but  readily  in  ether 
or  hot  alcohol.  Solanidine  is  a  stronger  base  than  solanine  and  the  salts  it  forms 
with  acids  are  usually  crystalUne  and  difficulty  soluble  in  water.  Solanine  and 
solanidine  are  highly  toxic  substances  having  an  action  similar  to  that  of  the 
saponin  substances  (see  above). 

Toxic  Action. — Solanine  taken  internally  is  usually  very  imperfectly  ab- 
sorbed. As  a  glucoside  its  action  is  local  and  as  a  saponin-hke  substance  strongly 
haemolytic,  rendering  the  blood  laky.  A  solanine  solution  even  in  a  dilution 
of  1 :  8300  causes  complete  haemolysis.  Internal  administration  of  solanine  usually 
produces  emesis  and  larger  doses  cause  gastro-enteritis  (gastro-intestinal  catarrh) . 
The  latter  also  follows  intravenous  and  subcutaneous  injection  of  doses  not  rapidly 
fatal.  At  the  same  time  a  hemoglobinuria  may  appear.  (R.  Robert,  In- 
toxikationen). 

Detection  of  Solanine  and  Solanidine 

Since  very  dilute  mineral  acids  hydrolyze  solanine,  these  acids 
cannot  be  used  to  detect  this  alkaloid.  E.  Schmidt^  suggests 
the  following  procedure.  Extract  the  material  with  cold  water 
containing  tartaric  acid.  Neutralize  the  filtered  extract  with 
calcined  magnesia  and  evaporate  to  dryness  upon  the  water- 
bath.     Extract  the  residue  with  alcohol  and  filter  hot.     If  the 

1  Pharmazeutische  Chemie,  Organischer  Teil. 


POISONS   NOT   IN    THE    THREE   MAIN   GROUPS  210 

quantity  of  solaninc  is  not  too  small,  the  alcoholic  extract  gel- 
atinizes upon  cooling.  Otherwise,  evaporate  the  alcoholic 
solution  and  examine  the  residue  for  solanine.  L.  Kobert 
extracts  solanine  with  isobutyl  alcohol  from  alkaline  solution. 
Phospho-molybdic  acid  is  the  only  general  reagent  giving  a  pre- 
cipitate with  a  solanine  solution  and  that  is  yellow.  But  sol- 
anidine,  that  is  to  say,  a  solanine  solution  that  has  been  boiled 
with  excess  of  hydrochloric  acid,  being  a  stronger  base,  gives 
precipitates  with  most  of  the  other  general  reagents. 

Special  Tests  for  Solanine  and  Solanidine 

1.  A  solution  of  solanine  in  selenic-sulphuric  acid^  is  rasp- 
berry red.  Gentle  heat  favors  the  appearance  of  this  color. 
Solanidine  gives  the  same  result. 

2.  Solutions  of  solanine  and  solanidine  in  vanadic-sulphuric 
acid^  are  orange-yellow,  soon  becoming  red  and  finally  blue- 
violet.  Solanine  may  be  dissolved  first  in  sulphuric  acid  and  a 
drop  of  vanadic-sulphuric  acid  added  to  this  solution. 

3.  Solutions  of  solanine  and  solanidine  in  ethyl-sulphuric  acid^ 
are  red.  An  alcoholic  solution  of  solanine,  carefully  added  to 
concentrated  sulphuric  acid  as  an  upper  layer,  produces  a  red 
zone  where  the  two  liquids  meet  (E.  Schmidt). 

4.  A  solution  of  solanine  in  concentrated  sulphuric  acid  is 
orange  but  becomes  brownish  red  on  longer  standing  or  gentle 
warming.  Red  streaks  appear,  if  bromine  water  is  added  drop 
by  drop  to  a  solution  of  solanine  in  concentrated  sulphuric 
acid. 

5.  A  solution  of  solanine  in  Froehde's  reagent  is  first  yellow- 
ish red  then  evanescent  cherry  red  and  finally  red-brown. 

The  methods  for  estimating  solanine  quantitatively  in  pota- 
toes are  described  in  Chapter  VI  (see  page  284). 

^  A  mixture  of  1.3  grams  of  sodium  selenate  (NaoSeO^.io  H2O),  S  cc.  of  water 
and  6  cc.  of  concentrated  sulphuric  acid. 

-  Dissolve  0.1  gram  of  ammonium  vanadate  (H4N.VO3)  in  100  grams  of  concen- 
trated sulphuric  acid. 

^  Add  6  cc.  of  concentrated  sulphuric  acid  to  9  cc.  of  absolute  alcohol. 


H     H 

C      C     N.CH3 

HC      C      CH  CH2 

CH3O.C      C      C      CH2 

C      C      CH 

1        '        ' 

0- 

1          1 

-CH  CH 

\/ 
C 

1 

0CH3 

R.  Pschorr's  formula 

220  DETECTION   OF   POISONS 

THEBAINE 

Thebaine,  C19H21NO3  =  Ci7Hi5(OCH3)2NO,  constitutes  about  0.15  per  cent, 
of  opium.  This  alkaloid  crystallizes  from  dilute  alcohol  in  leaflets  having  a 
silvery  glitter  and  from  absolute  alcohol  in  prisms 
melting  at  193°.  It  is  nearly  insoluble  in  water, 
rather  easily  soluble  in  hot  alcohol,  ether,  benzene 
and  chloroform.  It  differs  from  morphine  in  being 
nearly  insoluble  in  caustic  alkaUes.  Its  solutions  are 
tasteless  and  laevo-rotatory. 

Constitution. — Thebaine  is  a  strong  tertiary  base, 
forming  as  a  rule  well  crystallized  salts  with  acids. 
But  excess  of  acid,  especially  mineral  acid,  usually 
decomposes  these  salts  with  ease.  Being  a  tertiary 
base,  it  easily  combines  with  methyl  iodide,  forming 
thebaine  iodomethylate,  C19H21NO3.CH3I,  crystalliz- 
ing in  prisms.  Two  of  the  three  oxygen  atoms  in 
thebaine  are  methoxyl-groups  (  —  OCII3)  and  the  third  probably  forms  an 
ether-like  combination,  a  so-called  bridge-oxygen.  The  thebaine  molecule 
appears  not  to  contain  hydroxyl. 

Heated  with  acetic  anhydride,  thebaine  gives  the  acetyl  derivative  of  the 
phenol  thebaol,  C16H14O3,  and  a  nitrogenous  product,  methyl-oxy-ethylamine, 
CH3.NH.CH2.CH2.OH.  R.  Pschorr  has  synthesized  thebaol,  or  the  methyl 
ether  of  thebaol,  and  shown  by  this  synthesis  that  thebaol  is  3,6-dimethoxy- 
4-oxy-phenanthrene  (see  below).  Pschorr  assigns  to  thebaine  the  structural 
formula  given  above  which  is  analogous  to  that  of  apomorphine  and  of  morphine 
(see  pages  122  and  126).     Thebaol  has  the  following  structural  formula: 

(i)H     H 
C      C 

/\/\ 
HC      C      CH 

(3)CH30.C      C      C 

C      C       CH 

I        I         il 
(4)H0  HC       CH 

\/ 
C 

OCH3(6) 

Detection  of  Thebaine 

Ether  and  chloroform  extract  thebaine  from  an  alkaline 
aqueous  solution  and  consequently  this  alkaloid  appears  in 
ether  extract  B,  if  the  Stas-Otto  procedure  is  followed.  The 
general  reagents,  phospho-tungstic  acid,  iodo-potassium  iodide, 
potassium  mercuric  iodide  and  potassium  bismuthous  iodide 


POISONS    NOT   IN   THE    THREE    MAIN    GROUPS  221 

precipitate  thcbaine  even  from  very  dilute  solutions.     Thebaine 
gives  the  following  color  reactions: 

1.  Concentrated  Sulphuric  Acid  gives  a  deep  red  color  with 
thebaine  and  the  solution  gradually  becomes  yellowish  red. 
Froehde's  reagent  gives  the  same  result. 

2.  Concentrated  Nitric  Acid  dissolves  thebaine  with  a  yellow 
color.  With  Erdmann's  reagent  the  color  varies  from  dark  red 
to  orange. 

3.  Chlorine  Water  dissolves  thebaine  and  ammonia  turns  the 
solution  an  intense  red-brown. 

Toxalbumins 

Toxalbumins  are  toxic,  protein-like  substances  either  already  formed  in  the 
plant  or  animal  organism,  or  produced  in  the  metabolism  of  pathogenic  micro- 
organisms. These  substances  as  yet  have  not  been  isolated  pure  as  individual 
chemical  compounds.  The  chemical  and  physiological  .  properties  of  such 
vegetable  toxalbumins  as  abrin,  ricin,  robin  and  crotin  are  given  as  a  matter  of 
fact  by  substances  obtained  from  some  particular  part  of  the  plant  by  a  definite 
method.  The  vegetable  toxalbumins  mentioned  possess  the  common  property 
of  clumping,  agglutinating  and  precipitating  red  blood  corpuscles.  Therefore 
R.  Kobert  classifies  them  as  "vegetable  agglutinines."  A  trace  of  one  of  these 
agglutinines,  added  to  defibrinated  blood  in  a  test-tube,  causes  clumping  into  a 
mass  resembling  sealing-wax.  Abrin,  ricin  and  crotin  also  cause  coagulation  of 
milk. 

Abrin 

This  toxalbumin  occurs  in  jequirity  seeds  from  Abrus  precatorius.  Remove 
the  seed  envelopes  and  extract  the  finely  divided  seeds  with  4  per  cent,  sodium 
chloride  solution.  Concentrate  the  filtered  liquid  -in  vacuo  and  acidify  with 
acetic  acid.  Precipitate  abrin  from  this  solution  by  addition  of  sodium  chloride 
and  finally  purify  by  dialysis.  Abrin  is  an  amorphous,  highly  toxic  powder  not 
entirely  free  from  ash.  Though  abrin  and  ricin  are  alike  in  some  respects,  they 
are  not  identical. 

Ricin 

This  intensely  toxic  toxalbumin  constitutes  2.8-3  P^r  cent,  of  the  castor  bean. 
Remove  the  seed  envelopes  and  subject  the  seeds  to  powerful  pressure  to  remove 
as  much  oil  as  possible.  Then  extract  uith  10  per  cent,  sodium  chloride  solution. 
Saturate  the  filtered  extract  at  the  same  time  mth  magnesium  and  sodium  sul- 
phate and  keep  for  some  time  in  the  cold  at  room  temperature.  Place  the  pre- 
cipitate, which  contains  ricin,  in  a  parchment  paper  diah-zing  tube  and  dial>-ze 
for  several  days.     Finally  dry  the  ricin  left  ///  vacuo  over  sulphuric  acid.     Ricin  is 


222  DETECTION   OF  POISONS 

an  amorphous,  highly  toxic  powder  containing  ash  and  easily  soluble  in  lo  per 
cent,  sodium  chloride  solution.  This  toxalbumin,  dissolved  in  sodium  chloride 
solution,  gives  the  protein  reactions.  Ricin  possesses  in  high  degree  the  power  of 
agglutinating  blood  corpuscles.  Use  defibrinated  blood  for  this  test-tube  experi- 
ment, not  diluted  blood  or  blood  mixed  with  physiological  salt  solution.  Ricin, 
according  to  Elf  strand,  agglutinates  the  red  blood  corpuscles,  of  the  guinea-pig 
even  in  a  dilution  of  i  :  600,000.  Ricin  agglutinates  the  blood  of  all  mammals  but 
not  to  the  same  degree.  Removing  serum  from  the  blood  and  substituting 
physiological  salt  solution  strengthens  rather  than  weakens  the  agglutinating 
action  of  ricin.  The  inference  is  that  serum  must  have  a  certain  anti-agglu- 
tinating  action.  Separation  of  red  blood  corpuscles  into  stroma  and  h£emo- 
globin^  shows  that  ricin  has  not  changed  hemoglobin  in  the  least.  But  the 
strolnata  have  been  altered  just  as  the  blood  corpuscles  have  been. 

To  detect  ricin  in  castor  bean  press -cake,  or  in  feeds  containing  castor  beans, 
extract  the  finely  divided  material  with  physiological  salt  solution  at  room  tem- 
perature, filter  and  make  the  agglutination  test  in  a  test-tube  with  undiluted,  de- 
fibrinated blood  and  with  blood  diluted  with  physiological  salt  solution. 

Crotin 

Crotin  is  a  substance  obtained  from  the  seeds  of  Croton  Tiglium.  Remove  the 
seed  envelopes,  express  the  oil  and  treat  as  described  for  abrin  and  ricin.  Chem- 
ically crotin  is  very  similar  to  ricin.  Abrin  and  ricin  agglutinate  the  blood  cor- 
puscles of  all  warm-blooded  animals  thus  far  tested  but  crotin  does  not  behave 
the  same  with  all  kinds  of  blood.     (See  R.  Kobert,  Intoxikationen.) 

Coagulation  of  Blood  and  Defibrinated  Blood 

Blood  is  a  transparent  fluid,  the  blood  plasma,  suspended  in  which  is  a  very 
large  number  of  solid  particles,  the  red  and  white  blood  corpuscles.  Outside 
the  organism  blood  coagulates  even  in  a  few  minutes  after  being  drawn.  In  the 
clotting  of  blood  a  very  difficultly  soluble  protein,  called  fibrin,  separates.  If  the 
blood  is  still,  the  clot  is  a  solid  mass  which  gradually  contracts  and  exudes  a  clear 
liquid,  usually  yellow,  the  blood  serum.  The  coagulum,  thus  formed  and  envelop- 
ing the  blood  corpuscles,  is  called  the  crassamentum  (Placenta  sanguinis).  But 
if  the  blood  is  whipped  during  coagulation,  fibrin  separates  in  threads.  The 
fluid  separated  from  the  latter  is  defibrinated  blood  which  consists  of  blood  cor- 
puscles and  blood  serum.  To  obtain  defibrinated  blood,  whip  the  fresh  blood 
removed  from  a  vein  with  twigs  and  fibrin  will  separate  on  these.  Or  run  the 
fresh  blood  into  an  Erlenmeyer  flask,  containing  iron  filings,  and  shake  vigorously 
for  several  minutes.     Fibrin  is  precipitated  on  the  filings. 

There  are  several  ways  to  retard  coagulation  of  blood,  among  which  the  follow- 
ing may  be  mentioned: 

I.  Cool  blood  suddenly  to  low  temperature. 

^  The  two  principal  components  of  blood  corpuscles  are  the  stroma,  which  con- 
stitutes the  true  protoplasm,  and  the  intraglobular  contents,  the  chief  constituent 
of  which  is  haemoglobm. 


POISONS  NOT  IN  THE  THREE  MAIN  GROUPS      223 

2.  Draw  blood  direct  from  the  vein  into  a  natural  salt  solution,  for  cxamj^lc, 
magnesium  sulphate  solution  (i  volume  of  salt  solution  and  3  volumes  of  blood) 
and  stir.     This  mixture  of  blood  and  salt  will  not  coagulate  for  a  day. 

3.  Add  blood  to  sufficient  dilute  potassium  oxalate  solution  to  give  a  mixture 
containing  0.1  per  cent,  of  oxalate.  The  soluble  calcium  salts  of  the  blood  are 
precipitated  by  the  oxalate  and  the  blood  loses  its  power  of  coagulating. 

4.  To  prepare  a  non-coagulating  blood  plasma,  pour  blood  into  sodium  fluoride 
solution  until  it  contains  0.3  per  cent,  of  NaF. 


CHAPTER  V 

SPECIAL  QUALITATIVE  AND  QUANTITATIVE  METHODS 

Quantitative  Estimation  of  Phosphorus  in  Phosphorated  Oils 

I.  W.  Straub's  Method. — Straub  has  found  that  his  test^  with 
dilute  copper  sulphate  solution,  recommended  for  the  qualita- 
tive detection  of  phosphorus,  may  also  be  used  to  determine 
phosphorus  in  a  phosphorated  oil.  If  such  an  oil  is  shaken  with 
3  per  cent,  copper  sulphate  solution,  there  is  first  a  brownish 
black  emulsion  in  which  each  individual  oil  drop  is  coated  with  a 
film  of  copper  phosphide,  PCus  (?).  After  4-5  hours  shaking, 
this  brownish  black  color  disappears  and  the  mixture  separates 
into  two  layers.  All  the  phosphorus  in  the  oil  is  now  in  the 
aqueous  solution  as  phosphoric  acid.  This  method  has  the 
further  advantage  that  the  decolorization  of  the  emulsion  serves 
as  an  indicator  of  the  completion  of  the  oxidation. 

Procedure. — Put  25  cc.  of  3  per  cent,  copper  sulphate  solution 
(taken  as  CUSO4.5H2O)  in  a  separatory  funnel.  Add  5  cc.  of 
the  phosphorated  oiP  and  agitate  the  mixture  vigorously  for  a 
long  time.  If  a  shaking  machine  is  available,  place  the  mixture 
in  a  thick-walled  glass  bottle  with  a  tight  glass  stopper  and  shake 
3-5  hours,  or  until  the  original  brown  emulsion  has  disappeared 
and  become  clear  and  bright  blue.  Separate  the  aqueous  solu- 
tion in  a  separatory  funnel  and  precipitate  phosphoric  acid  at 
once  by  the  molybdate  method  and  finally  weigh  as  magnesium 
pyrophosphate,  Mg2P207. 

^  Zeitschrift  fiir  anorganische  Chemie  35,  460  (1903). 

^  To  prepare  a  phosphorated  oil  suitable  for  such  determinations,  dissolve  about 
0.1  gram  of  yellow  phosphorus  in  the  smallest  possible  quantity  of  warm  carbon 
disulphide  and  dilute  this  solution  to  100  cc.  with  olive  oil.  Although  carbon 
disulphide  does  not  affect  the  determination  of  phosphorus,  it  may  be  removed 
by  warming  the  phosphorated  oil  on  the  water-bath. 

[224 


SPECIAL  QUALITATIVE    AND   QUANTITATIVE    ME'IIKJDS       225 

Remarks. — The  accuracy  of  this  method  is  shown  by  the  results  of  Straub's 
determinations.  Instead  of  0.005  gram  of  phosphorus,  dissolved  in  5  cc.  of  oil, 
he  found  0.0047  and  0.00468  gram.  Even  very  considerable  dilutions  of  the 
phosphorated  oil  do  not  affect  the  accuracy  of  the  determination.  In  the  case  of 
the  more  concentrated  phosphorated  oils,  shaking  with  copper  sulphate  solution 
must  be  kept  up  much  longer. 

2.  A.  Frankel's^  and  C.  Stich's"  Method. — Dissolve  the  oil 
in  acetone  and  precipitate  phosphorus  with  hot  alcoholic  silver 
nitrate  solution.  Oxidize  the  phosphorus  in  the  precipitate  to 
phosphoric  acid  and  finally  determine  the  latter  in  the  usual 
way. 

Procedure. — Dissolve  20-50  cc.  of  the  phosphorated  oil,  as 
phosphorated  cod  liver  oil,  in  100  cc.  of  acetone  or  ether  and 
completely  precipitate  with  hot  alcoholic  silver  nitrate  solu- 
tion.' First  wash  the  precipitate  of  silver  phosphide  with 
ether-acetone  mixture  and  then  with  alcohol.  Treat  next  with 
hot  25  per  cent,  nitric  acid,  containing  a  Httle  fuming  acid. 
Expel  excess  of  nitric  acid  from  the  filtrate  on  the  water-bath 
and  precipitate  silver  with  hydrochloric  acid.  Finally  filter 
from  silver  chloride  and  determine  phosphoric  acid  in  the  filtrate. 

Remarks. — Since  sodium  hypophosphite  and  phosphite  are  soluble  in  acetone 
and  also  precipitated  by  acetone-silver  nitrate,  it  is  advisable  first  to  extract  a 
test  portion  of  the  phosphorated  oil  with  water  and  then  test  the  aqueous  extract 
for  these  first  oxidation  products  of  phosphorus,  h5^ophosphorous  and  phosphor- 
ous acids.  If  they  are  present,  all  the  phosphorated  oil  should  first  be  extracted 
with  water  in  the  same  manner. 

Phosphorus  in  phosphorated  oils,  especially  phosphorated  cod  liver  oil  slowly 
disappears.  C.  Stich  found  that  a  phosphorated  cod  liver  oil,  containing  0.05 
per  cent,  of  phosphorus,  with  the  usual  daily  removal  of  5  grams,  lost  in  3  weeks 
only  3-5  milligrams  of  phosphorus.  Such  a  decrease  in  the  amount  of  phosphorus 
in  phosphorated  oils  is  only  of  slight  significance.  Dilute  oily  solutions  of  phos- 
phorus (i:  1000),  when  kept  in  tightly  stoppered  bottles  and  protected  from  light, 
are  constant  as  regards  their  phosphorus  content  for  a  long  time,  even  5-6  months. 
Moreover,  phosphorous  much  diluted  as  vapor  or  in  solution,  is  oxidized  ^^•ith 
corresponding  difficulty.  The  same  is  also  true  of  phosphorus  in  the  animal  or- 
ganism. Therefore  it  is  possible  sometimes  to  detect  free  phosphorus  in  the 
excretory  organs,  as  the  liver,  even  several  weeks  after  phosphorus  posioning. 

The  distillation  method  is  inapplicable  in  the  quantitative  estimation  of  phos- 

^  Pharmazeutische  Post  34,  117. 
2  Pharmazeutische  Zeitung  37,  500  (1902). 
^  Silver  nitrate  dissolves  in  about  10  parts  of  alcohol. 
15 


226  DETECTION    OF   POISONS 

phorus  in  oils,  as  cod  liver  oil,  since  only  about  40  per  cent,  of  the  phosphorus 
present  is  fourid  in  the  receiver,  even  when  the  strongest  oxidizing  agent  and  the 
best  absorbent  for  phosphorus  are  used.  To  place  the  phosphorus-content  of  the 
cod  liver  oil  residue  at  the  amount  of  the  distilled  phosphorus  is  not  admissible, 
because  cod  liver  oil  as  such  contains  about  0.02  per  cent,  of  combined  phosphorus. 

Special  Methods  for  the  Detection  of  Arsenic 
Isolation  of  Arsenic  as  Arsenic  Trichloride^ 

This  depends  upon  the  volatility  of  arsenic  as  chloride,  AsCls, 
in  concentrated  hydrochloric  acid  solution  and  in  presence  of 
ferrous  chloride.  The  latter  serves  (a)  to  reduce  any  arsenic 
acid  possibly  present  in  the  material  to  arsenious  acid  which  with 
concentrated  hydrochloric  acid  then  forms  arsenic  trichloride 

(a)  H3ASO4  +  2HCI  +  2FeCl2  =  H3ASO3  +  H2O  +  2FeCl3, 
03)  H3ASO3  +     3HCI  =  ASCI3      +  3H2O. 

Procedure. — Comminute  the  material  and  mix  with  very 
concentrated  hydrochloric  acid  (about  40  per  cent.)  until  rather 
thin.  Then  add  5  grams  of  20  per  cent,  arsenic-free  ferrous 
chloride  solution  or  saturated  ferrous  sulphate  solution  and 
put  the  mixture  into  a  capacious  retort,  the  neck  of  which  is 
directed  obliquely  upward  and  connected  with  a  Liebig  cooler 
by  an  obtuse  angle  tube,  and  carefully  distil.  Distil  about  a 
third  to  a  half  of  the  original  mixture.  Dilute  the  distillate 
with  water  and  test  for  arsenic  in  the  Marsh  apparatus,  using 
hydrochloric  acid  for  the  evolution  of  hydrogen. 

If  a  tubulated  retort  is  used  for  the  distillation,  hydrochloric  acid  gas  can  be 
passed  in  during  distillation  so  that  the  liquid  being  distilled  is  kept  saturated  with 
this  acid. 

Electrolytic  Detection  of  Arsenic 

To  detect  arsenic  electrolytically,  put  the  liquid,  as  the  sul- 
phuric acid  solution  obtained  according  to  the  general  procedure 
which  contains  arsenic  as  arsenic  acid_^(see  page  150),  or  urine  or 
stomach  contents,  in  a  sufficiently  wide  U-tube  with  platinum 
electrodes    (Fig.    18).     Pass   the   current   through   the   Hquid 

^  H.  Beckurts,  Archiv  der  Pharmazie  222,  653  (i{ 


SPECIAL   QUAl.irA'I'IVK    AND    QUANTI'I'A'IIVE    METHODS       227 

acidified  with  sulphuric  acid,  and  arsinc,  AsHg,  together  with 
hydrogen  will  appear  at  the  cathode,  if  the  liquid  contains  ar- 
senic. First  test  the  hydrogen  for  arsenic  by  the  Gutzeit  arsenic 
test  (see  page  1 56) .  If  a  yellow  spot  appears  on  the  paper  moist- 
ened with  saturated  silver  nitrate  solution,  arsenic  is  present. 
That  this  is  actually  arsenic  may  be  shown  by  connecting  the  U- 
tube  as  shown  in  the  sketch  with  a  chloride  of  calcium  tube  and 
a  Marsh  reduction  tube;  an  arsenic  mirror  then  appears  in 


Fig.  18. — Apparatus  for  the  Electrolytic  Detection  of  Arsenic. 

the  latter  when  heated  to  redness.  Use  a  current  having  an 
electromotive  force  of  7-8  volts.  The  electrolytic  method  is 
especially  adapted  for  the  detection  of  arsenic  in  inorganic 
compounds  present  in  secretions,  as  the  urine,  but  not  for  arsenic 
in  organic  combination  as  cacodyl  compounds  and  arrhenal.  An 
exception  among  these  organic  compounds  of  arsenic  is  atoxyl, 
or  the  anihd  of  meta-arsenic  acid,  AsO2.NH.C6H5.  The  arsenic 
being  rather  loosely  bound  is  broken  up  by  the  electric  current 
with  formation  of  arsine. 

Destruction  of  Organic  Matter  and  Detection  of  Arsenic 
(According  to  A.  Gautier^  and  G.  Lockemann-) 

This  method  is  of  scientific  interest  rather  than  of  practical  significance  in 

^  Bulletin  de  la  Societe  chimique  de  Paris,  29,  639  (1903). 

^  Zeitschrift  fiir  angewandte  Cheniie  18,  416,  491  (1905);  also  19,  1362  (1906). 


228  DETECTION    OF    POISONS 

forensic  chemistry.  The  purpose  is  to  increase  the  delicacy  of  the  Marsh- 
BerzeUus  test  for  arsenic,  and  to  exclude  as  far  as  possible  sources  of  error  con- 
nected with  the  destruction  of  organic  matter,  the  precipitation  of  arsenic  with 
hydrogen  sulphide  and  the  evolution  and  drying  of  hydrogen  gas.  Organic 
matter  is  destroyed  without  the  use  of  hydrochloric  acid,  and  arsenic  is  detected 
without  precipitation  as  arsenic  sulphide.  Lockemann  recommends  the  following 
procedure  and  uses  finely  divided  meat  as  a  test  experiment: 

Place  20  grams  of  finely  chopped  meat  in  a  porcelain  dish  and  add  a  few  cc. 
of  a  mixture  to  10  parts  of  fuming  nitric  acid  and  i  part  of  concentrated  sulphuric 
acid.  Warm  upon  the  water-bath.  The  action  of  the  acid  mixture  is  so  vigorous 
that,  even  after  the  addition  of  about  5  cc,  the  entire  mass,  which  puffs  up  con- 
siderably at  first,  changes  to  a  yellowish,  homogeneous,  thick,  oily  liquid.  If  too 
much  acid  is  added  at  once  during  warming  upon  the  water-bath,  the  action  may 
be  violent  enough  to  cause  sudden  charring  of  the  whole  mass  with  copious  evolu- 
tion of  smoke.  Such  an  occurrence  may  result  in  loss  of  arsenic.  Consequently,  it 
is  advisable  to  add  the  acid  mixture,  amounting  in  all  to  about  20  cc,  to  the  meat 
in  1-2  cc.  portions,  not  adding  a  fresh  portion  of  acid  until  brown  fumes  cease 
coming  off.  The  mass  is  dark  yellow  and  finally  becomes  brown  after  long  heat- 
ing upon  the  water-bath.  Stir  with  a  concentrated  aqueous  solution  of  20  grams 
of  a  mixture  of  potassium  and  sodium  nitrate  (i  -f  i)  and  evaporate  upon 
the  water-bath.  There  remains  a  yellow,  crystalline  residue  which  still  contains 
organic  matter.  Gradually  introduce  this  mixture  in  small  portions  into  a 
platinum  crucible  containing  10  grams  of  fused  potassium  and  sodium  nitrate 
(i  -J-  i).  Having  added  all  the  mixture,  heat  the  crucible  for  a  short  time  over 
a  free  flame.  Dissolve  the  cold  melt  in  water,  add  sulphuric  acid  and  heat  upon 
the  water-bath  until  nitrous  fumes  have  been  expelled.  Test  a  cold  solution  of 
the  residue  for  arsenic  in  the  Marsh  apparatus. 

Lockemann  formerly  precipitated  arsenic  with  aluminium  hydroxide,  A1(0H)3. 
Add  10  cc.  of  a  1 2  per  cent,  solution  of  crystallized  aluminium  sulphate,  Al2(S04)3- 
18H2O  to  the  solution  of  the  melt  free  from  carbon  dioxide  and  nitrous  acid. 
Render  the  solution  alkaline  with  ammonia  and  heat  about  30  minutes  upon  the 
water-bath.  Collect  the  precipitate  upon  a  paper,  wash  with  water  containing 
ammonia  and  dissolve  in  about  30  cc.  of  10  per  cent,  sulphuric  acid.  Heat  the 
solution  in  a  porcelain  dish  upon  the  water-bath  until  it  no  longer  gives  a  test  for 
nitric  acid  with  diphenylamine-sulphuric  acid.^  Then  examine  this  solution  for 
arsenic  in  the  modified  Marsh  apparatus^  devised  by  Lockemann  (Fig.  19). 

Lockemann's  latest  results  have  shown  that  ferric  hydroxide  is  much  more 
effective  than  aluminium  hydroxide  as  a  precipitant  of  small  quantities  of  arsenic. 
Render  the  water  solution  of  the  melt  (see  above)  slightly  acid  with  sulphuric 
acid,  add  a  few  cc.  of  iron  alum  solution,  then  in  the  cold,  best  after  cooling  with 
ice,  add  just  enough  ammonia  to  precipitate  all  the  iron.  Filter  after  30  minutes, 
wash  the  precipitate  with  cold  water  to  remove  nitrates  completely,  then  dissolve 

1  Dissolve  I  gram  of  diphenylamine  in  100  grams  of  concentrated  sulphuric  acid. 
A  drop  of  the  liquid  with  a  drop  of  this  diphenylamine  solution  in  a  porcelain  dish 
should  not  give  a  blue  color. 

^  O.  Pressler,  30  Bruederstrasse,  Leipzig,  Germany,  supplies  this  apparatus  and 
also  the  ignition  tubes. 


SPECIAL   QUALITA'IIVK    AND    (^UAN'I  ITATI VK    MK'IIIODS 


229 


in  dilute  sulphuric  add  and  test  the  Kolution  for  arsenic  in  the  Marsh  api>aratus. 
Iron  salts  do  not  interfere  with  the  delicacy  of  the  Marsh  test  for  arsenic. 

Zinc  in  sticks^  and  sulphuric  acid  are  used  in  the  preparation  of  hydrogen. 
Copper  is  the  best  activator  of  zinc  in  the  Marsh  apparatus.  Break  the  zinc 
sticks  into  pieces  weighing  about  1.2-1.8  grams,  place  for  a  minute  in  0.5  per  cent, 
copper  sulphate  solution,  wash  with  water,  dry  with  filter  paper  and  preserve  care- 
fully in  a  closed  bottle.  This  procedure  does  not  interfere  with  the  formation  of 
the  mirror,  whereas  addition  of  copper  sulphate  to  the  reduction  flask  causes  re- 
tention of  arsenic.     Copper  sulphate  used  for  this  purpose  should  be  carefully 


Fig.  19. — Marsh  Apparatus  Modified  by  Lockemann. 


purified  by  several  recrystallizations.  The  basic  properties  of  fused  and  granu- 
lated calcium  chloride,  which  are  not  entirely  removed  even  by  hydrogen  chloride 
and  carbon  dioxide,  make  this  an  unsuitable  drying  agent  for  hydrogen.  Locke- 
mann found  that  potassium  carbonate,  phosphorus  pentoxide  and  concentrated 
sulphuric  acid  cause  a  noticeable  decomposition  of  arsine,  and  the  same  is  true  of 
glass  wool  and  cotton.  Crystallized  calcium  chloride  in  pieces  about  i  cc.  in 
volume  is  the  best  drying  agent,  because  it  is  entirely  indifferent  to  arsine.  Locke- 
mann's  special  drying  tube  (see  sketch)  is  adapted  for  the  use  of  this  substance. 
Bohemian  glass,  having  a  wall  thickness  of  i  mm.  and  an  internal  diameter  of 
4  mm.,  is  used  for  ignition  tubes.  These  are  drawn  out  in  two  places  to  a  length  of 
4  cm.  The  outer  diameter  of  the  constriction  is  1.5  mm.  and  the  inner  about  0.5 
mm.  The  reduction  flask  contains  4-6  pieces  of  coppered  zinc  and  about  1 5  cc. 
of  15  per  cent,  sulphuric  acid  are  added  from  the  dropping  funnel.  After  hydro- 
gen has  been  passing  through  the  apparatus  for  30  minutes,  heat  is  apphed  in  front 

1  Lockemann  has   found   Kahlbaum's   stick   zinc   always   arsenic-free.     The 
same  may  be  said  of  Bertha  spelter  from  the  New  Jersey  Zinc  Companj'. 


230  DETECTION   OF   POISONS 

of  the  first  constriction  of  the  ignition  tube.  If  the  materials  are  arsenic-free 
after  1.5-2  hours  heating,  place  the  flame  in  front  of  the  second  constriction 
of  the  ignition  tube.  The  solution  of  the  iron  hydroxide  precipitate,  prepared  as 
described  above,  is  added  to  the  reduction  flask  from  the  dropping  funnel  which  is 
washed  with  a  little  water  or  dilute  sulphuric  acid.  In  testing  for  very  small 
quantities  of  arsenic,  it  is  advisable  to  cool  the  place  where  the  mirror  is  deposited 
by  keeping  the  cotton  thread  wet  (see  sketch). 

By  means  of  the  apparatus  described  Lockemann  has  detected  even  0.000 1  mg. 
of  arsenic  distinctly. 

Moist  air  gradually  oxidizes  the  arsenic  mirror,  but  in  an  absolutely  dry  at- 
mosphere even  when  exposed  to  light  there  is  no  change.  In  a  closed  tube  con- 
taining a  little  phosphorus  pentoxide  arsenic  mirrors  may  be  kept  unchanged  even 
for  months. 

Glass  wool,  or  cotton,  noticeably  decomposes  arsine.  The  decomposition  of 
arsine  in  aqueous  solution  is  also  hastened  by  the  presence  of  fine  filamentary 
bodies.     This  reaction  is  probably  catalytic  in  character. 

Electrolytic  Estimation  of  Minute  Quantities  of  Arsenic 

(C.  Mai  and  H.  Hurt') 

By  this  method  minute  amounts  of  arsenic  (fractions  of  a 
milKgram)  are  separated  quantitatively  at  the  cathode  from  an 
arsenical  electrolyte  as  arsine.  The  latter  then  reacts  quanti- 
tatively with  silver  nitrate  as  follows : 

AsHs  +  3H2O  +  6AgN03  =  H3ASO3  +  6HNO3  +  6Ag. 

The  advantages  of  the  electrolytic  detection  of  arsenic  are 
first  the  avoidance  of  traces  of  arsenic  that  sometimes  come 
from  zinc  in  the  Marsh  test  and  second  that  destruction  of  or- 
ganic matter  is  often  unnecessary.  T.  E.  Thorpe^  has  shown  the 
latter  to  be  the  case  in  the  examination  of  beer  worts  and  malt 
extracts  for  arsenic.  To  reduce  arsenic  acid  and  its  salts,  a  few 
drops  of  zinc  sulphate  solution  should  be  added  to  the  sulphuric 
acid  acting  as  the  electrolyte.  The  cathode  is  said  to  have  a 
higher  tension  and  the  hydrogen  to  be  very  active. 

Apparatus  and  Procedure. — The  apparatus  used  by  Mai  and 
Hurt  is  shown  in  Fig.  20. 

A  is  the  reduction  tube  and  B  a  bulb  tube  with  5-6  bulbs  con- 

^  Zeitschrift  fiir  Untersuchung  der  Nahrungs-  und  Genussmittel  9,  193  (1905) 
and  also  Pharmazeutische  Zeitung,  1905. 

^  Proceedings  of  the  Chemical  Society  19,  183  (1903). 


SPECIAL  QUALITATIVE    AND   QUANTITATIVE    ME'lHODS       231 


taining  o.oi  n-silver  nitrate  solution.  A  and  B  arc  connected 
by  a  small  tube  g  containing  pieces  of  pumice  stone  saturated 
with  an  alkaline  lead  solution,  or  glass  wool,  to  retain  any  traces 
of  hydrogen  sulphide.  Anode  a  and  cathode  e  are  lead  strips 
about  1-2  mm.  thick.  Their  upper  ends  about  5  mm.  thick 
are  luted  into  glass  tubes  b  which  pass  through  the  stopper  of  the 
U-tube  and  are  tight.     The  dropping  funnel  d  holds  about  25  cc. 


^=C=I==>. 


Fig.  20i — Apparatus  for  the  Electrolytic  Estimation  of  Arsenic. 

and  its  capillary  end  dips  about  2  cm.  into  the  solution  to  be 
electrolyzed.  Tube  c  for  the  escape  of  oxygen  from  the  anode 
chamber  contains  a  little  water. 

Fill  U-tube  A  up  to  the  mark  with  12  per  cent,  arsenic-free 
sulphuric  acid  and  bulb  tube  B  with  10  cc.  of  o.oi  n-silver  nitrate 
solution.  Turn  on  the  current  and  keep  at  2-3  amperes.  If 
the  silver  nitrate  solution  remains  unchanged  after  hydrogen 
has  been  running  i  hour,  the  lead  cathode  and  sulphuric  acid 
are  arsenic-free.  Without  stopping  the  current,  introduce  from 
the  dropping  funnel  the  solution  to  be  tested  for  arsenic,  the 
quantity  of  which  should  not  be  more  than  10  cc.  Add  this 
solution  as  slowly  as  possible  and  wash  the  last  traces  in  with  a 


232  DETECTION   OF  POISONS 

little  water.     If  the  solution  contains  arsenic,  or  arsenic  acid, 

the  silver  nitrate  solution  will  become  dark  in  a  few  minutes  and 

the  reaction  will  be  at  an  end  in  3  hours.     Pour  the  contents  of 

the  bulb  tube  through  a  small  asbestos  filter,  wash  with  3-4  cc. 

of  water  and  titrate  excess  of  o.oi  n-silver  nitrate  with  o.oi 

n-potassium  sulphocyanate  according  to  Volhard's  method. 

Calculation. — The  reaction  above  shows  that  6  molecules  of 

silver  nitrate  correspond  to  i  atom  of  arsenic  ( =  75) .     Therefore 

I  gram-molecule  of  silver  nitrate  =1/6  gram-atom  of  arsenic  = 

75 

-7-  =  12.5  grams  of  arsenic  and  1000  cc.  of  o.oi  n-silver  nitrate 

=  0.125  gram  of  arsenic. 

Notes. — Electrodes  of  platinum  (foil  or  gauze)  cannot  be  used  in  the  electro- 
lytic separation  of  arsenic  as  arsine,  because  either  solid  arsine  or  elementary 
arsenic  is  formed.  Mai  and  Hurt  also  found  that  gold,  silver  and  tin  cathodes 
gave  unsatisfactory  results  and  carbon  electrodes  were  not  much  better.  Pure 
lead  alone  meets  all  the  requirements  as  a  material  for  the  electrodes.  Oxygen 
compounds  of  arsenic  are  quickly  and  completely  reduced  to  gaseous  arsine  only 
upon  cathodes  of  absolutely  pure  lead.  The  attachment  of  a  platinum  wire  to  a 
lead  electrode  was  sufficient  to  cause  incomplete  reduction  of  arsenic  compounds. 
For  this  reason  the  electrodes  consist  of  one  piece  of  lead^  without  soldering  on 
wire  of  another  metal.  The  best  electrolyte  is  12  percent,  sulphuric  acid.  A 
stronger  acid  easily  causes  the  formation  of  hydrogen  sulphide  and  a  weaker  acid 
has  the  disadvantage  of  lower  conductivity  and  lower  specific  gravity.  The 
electrolyte  should  be  specifically  heavier  than  the  solution  to  be  tested  to  keep 
the  latter  from  passing  at  once  to  the  bottom  of  the  reduction  tube. 

Mai  and  Hurt  found  the  following  amounts  of  As: 

As  found 
0.223  mg. 
0.099  mg. 
0.105  mg. 

For  qualitative  tests  the  bulb  tube  may  be  replaced  by  the 
drying  and  ignition  tubes  of  the  Marsh  apparatus. 

According  to  Mai  and  Hurt  the  statements  of  Thorpe  and 
Trotmann,  that  every  solution  can  be  electrolyzed  without  pre- 
viously destroying  organic  matter,  do  not  always  hold.  In  the 
examination  of  beer  containing  arsenic  the  results  were  fairly 
satisfactory,  but  in  the  case  of  urine  the  results  were  far  too  high. 

^  Kahlbaum's  purest  lead. 


As  taken 

AS203 

0.25  mg. 

AS203 

o.io  mg. 

AS206 

o.io  mg, 

SPECIAL  QUALITATIVE    AND   QUANTITATIVK    METHODS 


233 


Quantitative  Estimation  of  Arsenic  and  Antimony  by  the  Gutzeit  Method 
Using  a  special  apparatus  and  paper  sensitized  with  mercuric 
chloride,  Sanger  and  Black^  have  found  that  the  Gutzeit  test 
can  be  employed  to  determine  small  amounts  of  arsenic  quanti- 
tatively. The  process  is  very  simple  and  requires  only  a  short 
time  for  completion.  Sanger  and 
RiegeP  have  extended  this  method 
to  the  estimation  of  antimony. 

Sensitized  Paper. — Paper  strips'' 
uniformly  4  mm.  wide  are  sensitized 
by  being  soaked  in  5  per  cent,  solu- 
tion of  recrystallized  mercuric  chlor- 
ide. These  are  dried,  cut  into  7  cm. 
lengths  and  protected  from  light  and 
moisture  in  a  stoppered  bottle  con- 
taining calcium  chloride,  or  soda 
lime,  covered  with  cotton. 
Apparatus. — A  30  cc.  bottle  (Fig. 

21)  for  the  reduction  is  closed  by  a 

glass  stopper  provided  both  with  a 

thistle  tube,  constricted  to  2  mm.  at 

the  end  and  extending  nearly  to  the 

bottom   of  the  bottle,  and  with  an 

exit  tube  widened  to  about  15  mm. 

just  above  the  stopper.     Connected 

with  this  exit  tube  by  a  ground  joint  and  at  a  right  angle  is  a 

tube  exactly  4  mm.  inside  diameter  and  approximately  9  cm.  in 

length  from  the  bend. 
Procedure. — Place  3  grams  of  uniformly  granulated  zinc^  in 

1  Proceedings  of  the  American  Academy  of  Arts  and  Sciences  43.  297-324 
(1907). 

2  Ibid.,  45,  21-27  (1909)- 

3  A  cold  pressed  paper  made  by  Whatman  has  been  found  to  give  the  best 
results. 

*  This  all  glass  apparatus,  suggested  by  ]Mr.  W.  A.  Boughton,  is  now  in  use  in  the 
Harvard  laboratory  and  is  a  modification  of  Sanger's  original  apparatus. 

6  Bertha  spelter  from  the  New  Jersey  Zinc  Company,  New  York,  has  been 
proved  free  from  arsenic. 


Fig.    21. — Apparatus   for   the 
Quantitative  Gutzeit  jMethod.* 


234  DETECTION   OF   POISONS 

the  bottle  and  a  strip  of  sensitized  paper  in  the  4  mm.  depositi6n 
tube.  In  estimating  arsenic  place  in  the  enlargement  of  the 
exit  tube  a  loose  plug  of  clean  absorbent  cotton  that  has  been 
kept  over  sulphuric  acid;  an  hour's  preliminary  run  is  necessary 
to  moisten  the  cotton  partially.  In  the  case  of  antimony  sub- 
stitute for  cotton  a  disc  of  filter  paper  that  has  been  moistened 
with  normal  lead  acetate,  dried  and  kept  in  a  well  stoppered 
bottle.  Before  inserting  this  disc  moisten  it  with  a  drop  of 
water.  Next  add  15  cc.  of  diluted  hydrochloric  acid^  (i  :6) 
and  let  the  hydrogen  run  10  minutes  to  make  sure  the  reagents 
cause  no  stain.  Then  add  the  whole,  or  an  aliquot  part  of  the 
solution  to  be  tested.  Arsenic  will  produce  a  color  on  the  paper 
in  a  few  minutes  which  will  reach  a  maximum  within  30  minutes. 
Antimony  produces  no  visible  effect  on  the  sensitized  paper, 
unless  the  amount  is  above  70  mmgr.  (  =  0.070  mg.)  when  a  gray 
color  may  appear.  If  there  is  any  color,  another  trial  should  be 
made  with  a  smaller  portion  of  solution.  In  the  determination 
of  arsenic  a  disc  of  lead  acetate  paper  should  be  inserted  beneath 
the  cotton  as  a  precaution  against  the  possible  formation  of 
hydrogen  sulphide. 

Standard  Bands. — (a)  Arsenic.  Dissolve  i  gram  of  resub- 
limed  arsenious  oxide  in  a  little  arsenic-free  sodium  hydroxide, 
acidify  with  sulphuric  acid  and  make  up  to  a  liter  with  recently 
boiled  water.  Dilute  10  cc.  of  this  solution  (I)  to  a  liter  with 
freshly  boiled  water  which  gives  a  solution  (II)  containing 
o.oi  mg.  of  arsenious-  oxide  per  cc.  Using  definite  volumes 
of  solution  II,  measured  from  a  burette,  prepare  a  series  of  color 
bands  (Fig.  22),  taking  a  fresh  charge  of  zinc  and  acid  for  each 
portion.  The  color  ranges  from  lemon  yellow  through  orange 
yellow  to    reddish    brown. 

(b)  Antimony. — Dissolve  2.3060  grams  of  pure,  recrystallized 
tartar  emetic  in  a  liter  of  water.  This  solution  (I)  contains  i.o 
mg.  of  antimonious  oxide  per  cc.  By  dilution  of  (I)  solutions 
containing  o.oi  mg.  (II)  and  o.ooi  mg.  (Ill)  are  prepared  and 
used  in  making  sensitized  bands. 

^  The  Baker  and  Adamson  Company  of  Easton,  Pennsylvania,  supply  a  very 
pure  acid  suitable  for  this  test. 


r 

■ 

i 

B 

1 

5  lo  IS  20  25  30  35  40  50  60  70 

Fig.  22. — Standard  Arsenic  Bands  in  Micromilligrams  of  AS2O3  (Initial). 


I 


5  10  IS  -o  ^S  30  35  40  50  'o  :d 

Tig.  22a. — Standard  Arsenic  Bands  in  Micromilligrams  of  AsoOs  (Hydrochloric  Acid 

Development) . 


mk 


-  5  10  15  20  25  30  35  40  52  :c  70 

Fig.  22b. — Standard  Arsenic  Bands  in  Micromilligrams  of  As^Os  (Ammonia  Development). 


SPECIAL   QUALITATJVK    AND   QUANTITA'llVE    METHODS       235 

These  bands  will  eventually  fade  but  they  may  be  preserved 
longer  by  being  sealed  in  glass  tubes  in  the  bottom  of  which  is 
phosphorus  pcntoxide  covered  with  cotton.  The  color  of  the 
arsenic  bands  may  be  developed  (i)  by  placing  the  band  in  hy- 
drochloric acid  (i  :  i)  for  2  minutes  at  a  temperature  not  over 
60°,  washing  thoroughly,  drying  and  sealing  as  before;  (2)  by 
treating  for  a  few  minutes  with  ammonium  hydroxide,  which 
gives  a  dense,  coal  black  color,  washing,  drying  and  seahng  in  a 
tube  over  quicklime. 

To  develop  the  antimony  band,  let  it  stand  in  a  test-tube 
covered  with  normal  ammonium  hydroxide  5  minutes.  A 
black  band  is  slowly  developed.  These  bands  may  be  protected 
as  described,  or  placed  between  glass  plates  cemented  together 
and  bound  with  passepartout  paper. 

The  more  dilute  standard  solutions  must  be  freshly  made  up 
within  a  few  hours  of  use. 

Notes. — Solutions  should  be  as  free  as  possible  from  sulphur  compounds 
jdelding  hydrogen  sulphide;  interfering  organic  matter;  and  metals  retarding 
formation  of  arsinc  and  stibine.  The  cotton  in  the  exit  tube  should  be  replaced 
after  10-12  runs,  and  the  lead  acetate  disc  after  each  run.  If  the  solution 
contains  arseniate,  reduce  with  10  cc.  of  arsenic-free  sulphurous  acid  and 
expel  the  excess. 

The  absolute  delicacy  of  the  method  is  set  at  0.00008  mg.  of  arsenious  oxide 
and  0.0005  n^g-  of  antimonious  oxide.  The  practical  delicacy,  using  a  band 
4  mm.  wide,  is  0.00 1  mg.  of  arsenious  oxide  and  0.002  or  0.003  ™g-  of  anti- 
monious oxide.  By  using,  however,  a  band  2  mm.  wide  in  a  correspondingly 
narrow  exit  tube,  a  practical  delicacy  of  0.0005  mg.  of  arsenious  oxide  and  o.oor 
mg.  of  antimonious  oxide  is  obtainable.  In  length  of  band  and  densitj'  of 
developed  color,  the  effect  of  arsine  on  the  sensitized  paper  is  from  2-3  times 
as  great  as  that  of  stibine.  The  authors  do  not  claim  a  greater  accurac}'  for  the 
method  than  within  10  per  cent. 

Biological  Detection  of  Arsenic  by  Penicillium  Brevicaiile 

B.  Gosio^  was  the  first  to  show  that  certain  moulds,  grown 
upon  media  containing  minute  quantities  of  arsenic,  produce 
volatile  arsenic  compounds  characterized  by  a  garlic-like 
odor.     Seven  species  of  moulds  were  found  to  have  this  power. 

^"Azione  di  alcune  muflfe  sui  composti  tissi  d'arsenico,"  Rivista  d'igiene  e 
sanita  publica,  1892,  201. 


236  DETECTION    OF   POISONS 

Penicillium  brevicaule,  however,  which  Gosio  isolated  from 
air,  and  which  was  first  found  upon  decaying  paper,  possessed 
this  property  in  the  highest  degree.  Gosio  states  that  we  are 
justified  in  regarding  PenicilHum  brevicaule  as  a  living  reagent 
for  arsenic.  Even  o.ooooi  gram  of  arsenic  can  be  recognized 
with  certainty  by  this  biological  test.  The  test  is  so  delicate 
that  it  should  be  of  great  value  in  toxicological  analysis  in  the 
preliminary  examination  for  arsenic. 

A.  Maassen^  states  that  a  temperature  of  28  to  32°  is  most 
favorable  to  the  growth  of  the  mould.  Crumbs  of  wheaten 
bread  were  found  to  make  an  especially  good  culture-medium. 
When  this  material  is  used,  a  vigorous  growth  of  mould  is 
visible  even  in  48  hours.  Sometimes  a  test  for  arsenic  can  be 
finished  in  a  few  hours,  and  always  in  2  or  3  days.  The  char- 
acteristic garlic  odor  from  weak,  arsenical  cultures  can  be  dis- 
tinctly recognized  even  after  several  months.  That  these 
"arsenic  moulds"  do  not  produce  gases  having  a  garlic  odor 
from  sulphur,  phosphorus,  antimony,  boron  and  bismuth  com- 
pounds, is  an  important  fact.  But  Penicillium  brevicaule 
possesses  in  high  degree  the  power  of  converting  solid  selenium 
and  tellurium  compounds  into  volatile  substances  having  a  pe- 
culiar odor.  The  odor,  especially  from  tellurium  cultures,  is 
like  that  produced  by  arsenic  cultures,  namely,  distinctly  like 
garlic!  The  odor  from  selenium  cultures,  however,  differs 
from  that  arising  from  arsenic  cultures.  It  is  more  of  a  mer- 
captan  odor. 

Biginelli^  found  that  the  gases,  generated  from  arsenic 
cultures  by  Penicillium  brevicaule,  are  completely  absorbed 
by  mercuric  diloride  solution.  Colorless  crystals,  having 
the  composition  (AsH(C2H5)2.2HgCl2),  are  formed.  This 
is  a  double  compound  of  mercuric  chloride  and  diethyl  arsine. 
This  compound  can  easily  be  decomposed.  It  then  diffuses 
an  intense  garlic  odor. 

R.  Abel  and  J.  Buttenberg^  state  that  a  mould  to  be  of 

^Arbeiten  aus  dem  Kaiserlichen  Gesundheitsamt,  1902,  478. 
^Chemisches  Centralblatt  (1900),  II,  1067,  and  also  (1900),  II,  iioo. 
^Zeitschrift  fur  Hygiene,  32,  440  (li 


SPECIAL   QUMJTATIVK    AND   QUANTITATIVI-:    METHODS       237 

use  in  the  biological  detection  of  arsenic  must  satisfy  the  fol- 
lowing conditions:  ''It  must  grow  rapidly,  and  not  generate 
any  odors  during  growth,  except  the  garlic  odor  produced  from 
an  arsenical  medium.  It  must  not  be  restricted  as  to  culture 
medium.  It  must  grow  in  presence  of  large,  or  very  small 
quantities  of  arsenic.  Finally,  it  must  demonstrate  its  specific 
action  in  presence  of  metallic  arsenic  and  all  kinds  of  arsenical 
compounds." 

The  best  material  for  these  experiments  is  white  or  Graham 
bread,  either  of  which  is  a  favorite  culture  tnedium  for  moulds. 
The  crust  is  the  only  part  of  bread  having  a  specific  aromatic 
odor.  When  this  has  been  removed,  the  crumbs  may  be  said 
to  be  practically  odorless. 

Procedure. — When  the  material  examined  is  Hquid,  absorb 
it  completely  by  adding  bread  crumbs,  and  scatter  a  small 
quantity  of  dry  bread  over  the  surface.  Solid  material  should 
be  finely  ground,  or  cut  into  as  small  pieces  as  possible,  and 
placed  in  not  too  small  a  flask.  Add  at  least  the  same  quantity 
of  bread  crumbs,  thoroughly  mix  the  two  substances  by  shaking, 
and  moisten  the  mass  with  a  little  water.  Close  the  flask  with 
a  cotton  plug,  and  sterilize  in  steam.  Sterilization  must  kill  all 
micro-organisms  in  the  flask.  Therefore,  heat  the  flask  in  an 
autoclave  lo  to  30  minutes  under  a  pressure  of  i  to  1.5  atmos- 
pheres. There  is  no  danger  of  volatizing  arsenic  during  ster- 
ilization. Then  inoculate  the  sterilized  material  when  cold. 
Place  in  a  flask  a  slice  of  potato,  superficially  coated  with 
mould  in  the  spore-forming  stage,  and  agitate  it  with  bouillon 
(peptone),  salt  solution  or  sterilized  water,  until  it  is  finely 
disintegrated.  Observe  all  necessary  precautions,  and  add 
the  mould,  suspended  in  water,  in  sufiicient  quantity  to 
impregnate  the  entire  surface  of  the  material  suspected  of 
containing  arsenic.  There  should  not  be  more  liquid,  how- 
ever, than  the  culture  medium  will  absorb.  Too  much  mois- 
ture retards  the  growth  of  the  mould.  Finally,  draw  a  tight 
rubber  cap  over  the  mouth  of  the  flask  and  cotton  plug.  Flasks 
thus  closed  may  stand  in  the  room,  but  it  is  better  to  keep  them 
at  a  higher  temperature,  for  example,  in  an  incubator  at  37°, 


238  DETECTION   OF   POISONS 

since  these  conditions  are  most  favorable  to  the  growth  of 
the  mould.  As  soon  as  a  growth  of  mould  is  distinctly  visible 
to  the  naked  eye  upon  the  medium,  the  first  indication  is  given 
that  a  test  of  the  culture  for  volatile  arsenic  compounds  may 
prove  stiiccessful.  In  a  very  favorable  case,  this  is  possible  in 
24  hours.  There  is  always  a  luxuriant  growth  of  mould  in  48 
to  72  hours,  so  that  a  decision  can  be  reached.  If  there  is  no 
odor,  the  flask  is  closed,  and  the  test  is  repeated  once  or  twice 
daily  on  the  following  days. 

Sulphuric,  hydrochloric  and  other  strong  mineral  acids  prevent  the  growth 
of  the  mould.  This  preventive  action  may  be  overcome  by  neutralization 
with  calcium  carbonate,  which  may  be  present  in  excess  without  ill  effect. 
Alkalies  also  interfere  with  the  growth  of  the  mould.  They  may  be  removed 
by  neutralization  with  tartaric  or  citric  acid,  either  of  which  may  be  present  in 
excess.  The  great  advantage  of  the  biological  over  the  purely  chemical  method 
lies  in  the  fact  that  less  time  is  required  to  get  a  result.  The  tedious  and  un- 
avoidable destruction  of  organic  matter  in  the  material  is  rendered  unnecessary. 
Moreover,  a  number  of  tests  for  arsenic  may  be  made  at  the  same  time. 

Abel  and  Buttenberg  (loc.  cit.)  speak  as  follows,  regarding  this  method: 
"The  biological  method  of  detecting  arsenic  has  so  many  advantages,  that  it 
deserves  to  be  recommended  for  the  most  varied  purposes.  Its  application  is 
very  general,  and  the  method  of  procedure  is  simple.  The  culture  of  the  mould 
can  be  kept  a  long  time,  even  a  year  or  more,  without  being  revived.  The 
test  is  very  delicate,  and  the  odor  is  readily  recognized.  The  generation  of  the 
odor,  in  the  case  of  cultures  containing  only  o.oooi  gram  of  arsenic,  can  be 
demonstrated  for  a  week." 

Besides  being  practically  unlimited  in  application,  the  biological  method  is 
extraordinarily  delicate.  In  this  respect,  it  exceeds  the  best  known  chemical 
methods  for  detecting  arsenic.  It  is,  for  example,  considerably  more  delicate 
than  Bettendorff's  test,  and  it  might  equal  in  delicacy  the  Marsh  and  Gutzeit 
tests. 

Detection  of  Arsenic  in  Organic  Arsenic  Compounds 
Cacodylic  Acid,  Arrhenal,  AtoxyP 

The  ordinary  reagents  usually  fail  to  show  arsenic  in  an  or- 
ganic arsenic  compound  dissolved  in  water.  Several  of  these 
compounds  persistently  resist  the  most  powerful  oxidizing  and 
reducing  agents. 

Cacodylic  Acid,  (CH3)2AsO-OH,  and  its  salts  have  been  used 
of  late  as  drugs.     A  2  per  cent,  solution  of  sodium  cacodylate, 

1  C.  E.  Carlson,  Zeitschrift  fiir  physiologische  Chemie  49,  410  (1906). 


SPECIAL   QUALITATIVE    AND   QUANTITATIVE    METHODS       230 

(CH3)2AsO-ONa.3H20,  conducts  the  electric  current  very  feebly 
but  no  arsine  appears  at  the  cathode.  Bettendorff's  reagent 
(stannous  chloride-hydrochloric  acid)  does  not  cause  separa- 
tion of  arsenic  from  cacodylic  acid  even  after  evaporation  with 
hydrochloric  acid  and  potassium  chlorate.  If  heated  with 
stannous  chloride-hydrochloric  acid,  cacodylic  acid  is  reduced 
to  the  foul  smelling  cacodylic  oxide,  [(CH3)2As]20,  recognized 
by  its  odor.  Distillation  of  sodium  cacodylate  by  Schneider's 
method  with  the  strongest  hydrochloric  acid  gives  no  arsenic 
trichloride  in  the  distillate.  The  arsenic  changes  to  another 
form,  not  precipitable  by  hydrogen  sulphide.  Evaporation  of 
the  distillate  upon  the  water  bath  with  nitric  acid  leaves  solid, 
non-volatile  cacodylic  acid  in  which  arsenic  may  be  detected  by 
reduction  with  sodium  carbonate-potassium  cyanide  mixture. 
Even  fuming  nitric  acid  does  not  oxidize  cacodylic  acid  to  ar- 
senious  or  arsenic  acid. 

Arrhenal,  Sodium  Methyl-Arseniate,  (CH3)AsO(ONa)2.5H20, 
forms  white  crystals  very  soluble  in  water.  Possibly  owing  to 
partial  hydrolysis,  an  aqueous  solution  of  this  compound  is 
alkaline  and  conducts  the  electric  current  feebly.  Only  traces 
of  arsine  appear  at  the  cathode  after  electrolysis  in  presence  of  a 
good  conductor.  In  arrhenal  the  arsenic  is  not  held  as  strongly 
as  in  the  cacodyl  compounds.  Hydrogen  sulphide  precipitates 
yellow  arsenic  trisulphide.  Distillation  with  strong  hydro- 
chloric acid  gives  arsenic  trichloride  in  the  distillate.  Betten- 
dorff's reagent  gives  a  red-brown  precipitate,  if  considerable 
arrhenal  is  present. 

Atoxyl,  the  Anilide  of  Metarsenic  Acid,  AsOo.NH.CeHs, 
forms  white,  odorless  crystals  readily  soluble  in  water  and  hav- 
ing a  faint,  saline  taste.  As  compared  with  cacodylic  acid, 
arsenic  in  atoxyl  is  less  firmly  bound.  Electrolysis  gives  arsine 
abundantly  at  the  cathode.  Hydrogen  sulphide  precipitates 
sulphide  of  arsenic.  Arsenic  trichloride  passes  over,  upon  dis- 
tillation wdth  concentrated  hydrochloric  acid.  Bettendorff's 
reagent  gives  a  lemon  yellow  precipitate. 

Urine. — In  suspected  arsenic  poisoning  first  examine  the  unne,  since  arsenic 
is  very  slowly  eliminated  bj^  this  channel.     Carlson  in  experiments  upon  him- 


240  DETECTION   OF  POISONS 

self  was  able  to  detect  arsenic  direct  in  the  urine  by  the  electrolytic  method  and 
also  by  the  Gutzeit  and  Marsh  tests.  He  took  lo  drops  of  Fowler's  solution^ 
daily.  Five  days  after  the  last  dose  Carlson  could  still  get  a  distinct  test  for 
arsenic  in  concentrated  urine.  The  urine  was  not  wholly  free  from  arsenic  until 
14  days  had  passed.  He  then  experimented  with  sodium  cacodylate,  taking 
daily  20  drops  of  a  i  per  cent,  solution.  He  could  not  detect  a  trace  of  arsenic 
in  the  urine  by  the  electrolytic  method.  Therefore  the  salt  of  cacodylic  acid 
had  passed  through  the  organism  unaltered.  But  cacodylic  acid  can  be  de- 
tected easily  in  the  urine,  upon  treating  the  latter  with  hypophosphorous  acid 
(sp.  gr.  1. 15). 2  Cacodylic  oxide  is  formed  and  can  be  recognized  by  its  odor. 
Sometimes  the  mixture  must  stand  several  hours  in  a  closed  test-tube.  Arrhenal, 
in  daily  doses  of  about  30  drops  of  i  per  cent,  solution,  behaved  like  the  cacodyl 
compound.  .Arsenic  could  not  be  detected  in  the  urine  by  electrolysis.  Con- 
sequently neither  arsenious  nor  arsenic  acid  had  been  formed  within  the  organism. 
Hypophosphorous  acid  immediately  precipitated  arsenic  from  arrhenal  and  gave 
the  cacodyl  odor. 

To  detect  cacodylic  acid  in  urine,  phosphorous  acid,  as  well  as  zinc  or  tin  and 
hydrochloric  acid,  may  be  used  instead  of  hypophosphorous  acid.  Frequently 
it  is  advisable  to  oxidize  most  of  the  organic  matter  in  the  urine  beforehand. 
Boil  25  cc.  of  urine  with  25  cc.  of  water,  5  per  cent,  potassium  permanganate 
solution  and  10  cc.  of  25  per  cent,  sodium  hydroxide  solution,  until  the  filtrate 
is  odorless  and  nearly  colorless.  Excess  of  hydrochloric  acid  (sp.  gr.  1.19)  and 
zinc  filings,  added  to  this  filtrate,  produce  with  heat  the  odor  of  cacodyl,  if  the 
urine  contains  cacodylic  acid.  Arsenic  from  atoxyl  can  be  isolated  at  the  cathode 
in  the  form  of  arsine  by  electrolysis.  Therefore  arsenic  can  be  detected  in  the 
urine  electrolytically  after  administration  of  atoxyl. 

Quantitative  Estimation  of  Minute  Amounts  of  Arsenic  -y^ 

(Karl  Th.  MornerO 

This  method  is  said  to  be  useful  in  estimating  arsenic  quantitatively  in  various 
kinds  of  fabrics  and  in  urine  in  cases  of  poisoning.  It  is  a  titration  method  de- 
vised especially  for  quantities  of  arsenic  not  exceeding  0.5  mg.  Arsenic 
is  first  precipitated  as  trisulphide  with  thioacetic  acid,  CH3.CO.SH.  Under  the 
conditions  arsenious  as  well  as  arsenic  acid  is  thus  precipitated.  In  alkaline  solu- 
tion potassium  permanganate  readily  oxidizes  arsenic  trisulphide  completely  to 
arsenic  acid  and  sulphuric  acid: 

AS2S3  -1-  14O  =  AszOs  +  3SO3. 

Potassium  permanganate  solution,  added  to  an  alkaline  solution  of  arsenic 
trisulphide,  immediately  loses  its  color,  being  decomposed  in  the  proportion  of 

^  Fowler's  solution  contains  i  per  cent,  of  AS2O3  as  potassium  arsenite. 

*  Instead  of  free  hypophosphorous  acid,  prepare  Engel  and  Bernard's  arsenic 
reagent  (Comptes  rend,  de  I'Acad.  des  sciences  122,  399),  or  J.  Bougault's  (I. 
Pharm.  Chim.  (6),  15,527).  Dissolve  20  grams  of  sodium  hj^pophosphite  in 
20  cc.  of  water  and  add  200  cc.  of  hydrochloric  acid  (sp.  gr.  1.17).  Filter  through 
a  cotton  plug  to  remove  NaCl  and  use  the  filtrate. 

'  Zeitschrift  fiir  analytische  Chemie  41,  397  (1902). 


SPECIAL  QUALITATIVE   AND   QUANTITATIVE   METHODS       241 

9  molecules  to  i  molecule  of  arsenic  trisulphide.'  Since  2  molecules  of  potassium 
permanganate  in  sulphuric  acid  solution  yield  5  atoms  of  oxygen  for  oxidation, 
9  molecules  according  to  the  proportion 

2:5  =  9:x     (x  =  22.5) 

should  give  22.5  atoms  of  oxygen.  But  according  to  the  reaction  above,  only 
14  atoms  of  oxygen  are  used  to  oxidize  i  molecule  of  arsenic  trisulphide,  whereas 
the  remaining  8.5  atoms  are  stored  up  in  the  precipitate  as  hydrated  manganese 
dioxide  (Mn02.H20).  But  if  the  reaction  mixture  is  heated  with  oxalic  acid  in 
presence  of  dilute  sulphuric  acid,  these  oxygen  atoms  become  active : 


(Mnl  O 


SO4  Hi 


O) >  CO:  OH 

"^  •  =  MnS04  +  H2O  +  2CO2  +  H2O. 


COO 


H 


Since  2  molecules  of  KMn04  yield  5  atoms  of  oxygen  and  since  14  atoms  of 
oxygen  are  necessary  for  i  molecule  of  AS2S3,  according  to  the  following  pro- 
portion 

Atoms      :    Mols.KMn04 

5  :  2  =14:  X     (x  =  S.6) 

5.6  molecules  of  potassium  permanganate  are  required  for  i  molecule  of  AS2S3 
(=  214),  or  2  atoms  of  arsenic  (=  150). 

1000  cc.  of  o.oi  n-potassium  permanganate  (=  0.3162  gram  KMn04)  contain 

in  solution r; — —  =  0.002  gram-molecule   of    KMn04  which  according 

10  X  10  X  10 

to  the  proportion 

Gram-mols.KMn04     :     Grm.  As 

5.6  :  150         =  0.002  :  X     (x  =  0.0536) 

represents  0.0536  gram  of  arsenic.  Hence  1000  cc.  of  o.oi  n-potassium  per- 
manganate solution  correspond  to  0.0536  gram  of  arsenic. 

Procedure. — Dissolve  arsenic  trisulphide  in  0.5  pei.  cent,  potassium  hydroxide 
solution^  and  run  this  solution  into  a  small  flask  containing  25  cc.  of  o.oi  n-potas- 
sium permanganate  solution.  Mix  the  contents  and  add  5  cc.  of  5  per  cent,  sul- 
phuric acid,  as  well  as  the  quantity  of  o.oi  n-oxalic  acid  solution  found  necessary 
by  special  titration.  Warm  until  the  color  is  discharged  and  finally  titrate  with 
O.OI  n-potassium  permanganate  solution. 

Preliminary  Titration. — Add  the  same  quantity  of  0.5  per  cent,  potassium 
hydroxide  solution  used  to  dissolve  arsenic  trisulphide,  as  well  as  5  cc.  of  5  per 
cent,  sulphuric  acid,  to  25  cc.  of  o.oi  n-potassium  permanganate  solution.  Heat 
the  mixture  to  boiling  and  add  oxalic  acid  solution  in  slight  excess  so  that  the 
liquid  becomes  colorless.     Titrate  back  with  o.oi  n-potassium  permanganate. 

^  Since  2  mols.  KMn04  give  in  alkaline  solution  3  atoms  of  available  oxj'gen 
(2KMn04  =  2Mn02  -f  3O  -|-  K2O),  i  mol.  of  AS2S3,  according  to  the  pro- 
Dortion:  3  :  2  =  14  :  x  (x  =  9.33),  requires  not  9  mols.  but  more  exactly  9.33 
mols.  of  KMn04. 

*  Ammonium   hydroxide   cannot   be   substituted   for   potassium   or   sodium 
hydroxide  solution. 
16 


242  DETECTION    OF   POISONS 

This  titration  shows  how  much  oxalic  acid  solution,  in  conjunction  with  traces 
of  reducing  substances  that  may  be  present  in  the  potassium  hydroxide  solution 
or  sulphuric  acid,  is  needed  in  a  regular  titration  for  the  exact  reduction  of  25 
c.c.  of  o.oi  n-potassium  permanganate. 

Example. — Suppose  that  25.5  cc.  of  oxalic  acid  solution  were  required  to  de- 
colorize the  boiling  liquid.  Titration  reqmred  0.3  cc.  of  o.oi  n-potassium  per- 
manganate solution.  Therefore  25  -f-  0.3  =  25.3  cc.  of  o.oi  n-potassium  per- 
manganate correspond  to  25.5  cc.  of  oxalic  acid  solution  and  25  cc.  of  the  former 
correspond  to  25.2  cc.  of  the  latter  solution.  Consequently  in  a  regular  titration 
25.2  cc.  of  O.OI  n-oxalic  acid  solution  must  be  used. 

The  amount  of  potassium  permanganate  in  25  cc.  of  o.oi  normal  solution  is 
sufl&cient  for  all  amounts  of  arsenic  up  to  0.5  mg.  Morner's  method  of 
determining  arsenic  gives  very  reliable  results  if  arsenic  is  in  the  form  of  the 
trisulphide  and  free  from  every  other  substance  soluble  in  0.5  percent,  potassium 
hydroxide  solution  and  capable  of  reducing  permanganate. 

Using  the  strongest  hydrochloric  acid,  Morner  first  distils  arsenic  as  arsenic 
trichloride  by  the  Schneider-Fife  method.  About  200  sq.  cm.  of  carpet,  100  sq. 
cm.  of  other  woven  and  paper  materials  and  15  grams  of  sealing  wax,  stearine 
or  wax  candles  and  dried  apples  were  used  for  each  determination  of  arsenic  by 
this  method.  According  to  Morner,  the  distillate  from  such  materials  by  the 
Schneider-Fife  method  always  contains  organic  matter,  even  when  caught  in 
dilute  nitric  acid.  To  remove  this  organic  matter  before  precipitating  arsenic 
with  thio-acetic  acid,  collect  the  distillate  in  a  receiver  containing  dilute  nitric 
acid  and  evaporate  to  dryness  in  a  porcelain  dish.  Add  to  the  small  residue  in  the 
dish  upon  the  water-bath  successively  2  cc.  of  potassium  hydroxide  solution 
(0.5  per  cent.  KOH)  heating  i  minute,  then  2  cc.  of  potassium  permanganate 
solution  (5  per  cent.  KMn04)  heating  about  3  minutes,  and  finally  i  cc.  of 
tartaric  acid  solution  (20  per  cent.  H2.C4H406)^  heating  until  the  color  is  dis- 
charged. Filter  into  a  porcelain  dish,  wash  the  filter  with  a  little  water  and  set 
the  dish  upon  a  boiling  water-bath.  Add  after  i  minute  i  cc.  of  thio-acetic  acid 
(5  per  cent.  CHs.COSH)^  and  warm  the  mixture  3  minutes.  Arsenic  is  precipi- 
tated as  arsenic  trisulphide.  After  cooling  for  5  minutes,  collect  the  precipitate 
upon  a  filter  and  wash  first  5  times  with  2  cc.  portions  of  0.5  per  cent,  sulphuric 
acid  and  then  3  times  with  2  cc.  portions  of  water.  Place  under  the  funnel  a 
small  flask  containing  25  cc.  of  o.oi  n-potassium  permanganate  solution  and  pour 
over  the  filter  3  portions  of  0.5  per  cent,  potassium  hydroxide  solution,  using  2  cc. 
each  time.  The  alkaline  solution  of  arsenic  trisulphide  thus  drops  directly  into 
the  permanganate  solution.  Otherwise,  proceed  as  described.  Subtract  0.3  cc. 
of  o.oi  n-permanganate  solution  from  the  volume  of  this  solution  used.     This 

^  Tartaric  acid  readily  dissolves  the  precipitate  of  manganese  peroxide.  To 
reduce  the  latter,  Morner  used  oxalic  and  lactic  acids,  sodium  sulphite  and  also 
thio-acetic  acid.  But  tartaric  acid  proved  to  be  better  than  any  of  these 
substances.  ^ 

^  Prepare  thio-acetic  acid  solution  by  shaking  5  cc.  of  thio-acetic  acid  with 
100  cc.  of  water.     Filter  and  keep  this  solution  in  a  dark  flask.     This  solution 
gradually  decomposes  with  evolution  of  hydrogen  sulphide: 
CH3.COSH  +  H2O  =  CH3.COOH  +  H2S. 


SPECIAL   QUALITATIVE    AND    QUANTITATIVE    METHODS        24 '> 

correction  is  necessary  because  even  the  finer  fjualitios  of  filter  paper  contain 
traces  of  substances  which  dissolve  in  0.5  per  cent,  potassium  hydroxide  solu- 
tion and  reduce  permanganate." 

Note. — The  procedure  described  separates  arsenic  trisulphide  from  every  other 
substance  soluble  in  0.5  per  cent,  potassium  hydroxide  solution  and  capable 
of  reducing  potassium  permanganate.  This  method  is  accurate  to  0.02  mg. 
of  arsenic. 

Detection  of  Salicylic  Acid  in  Foods  and  Beverages 

Wine.^ — Place  50  cc.  of  wine  in  a  cylindrical  separating  funnel 
with  50  cc.  of  a  mixture  of  equal  parts  of  ether  and  petroleum 
ether.  Shake  frequently,  taking  care  not  to  form  an  emulsion 
but  yet  to  mix  the  liquids  thoroughly.  Remove  the  ether- 
petroleum  ether  layer,  pour  through  a  dry  filter,  evaporate 
upon  the  water-bath  and  add  a  few  drops  of  ferric  chloride  solu- 
tion to  the  residue  which  becomes  red-violet  if  salicylic  acid  is 
present.  But  if  the  color  is  black  or  dark  brown,  add  a  few 
drops  of  hydrochloric  acid,  dissolve  in  water,  extract  with  ether- 
petroleum  ether  and  proceed  with  the  extract  as  just  described. 

Meat  and  Meat  Products.^ — For  experimental  purposes  add 
about  o.oi  gram  of  salicylic  acid  to  some  chopped  meat.  Ex- 
tract the  finely  divided  material  with  50  per  cent,  alcohol  and  add 
some  milk  of  lime  to  the  filtered  alcoholic  solution.  Evaporate 
to  dryness  upon  the  water-bath  and  stir  the  residue  with  a  shght 
excess  of  dilute  sulphuric  acid.  Shake  with  ether  without  fil- 
tering, pass  the  ether  extract  through  a  dry  filter  and  evaporate. 
Dissolve  the  residue  in  hot  water  and  test  the  filtered  solution 
with  very  dilute  ferric  chloride  solution  for  salicylic  acid. 

Milk."* — Mix  100  cc.  of  milk  with  100  cc.  of  water  at  60°. 
Precipitate  with  acetic  acid  and  mercuric  nitrate  solution,  using 
8  drops  of  each,  shake  and  filter.     Extract  the  filtrate  with  50 

^  After  passing  through  the  entire  process  in  several  blank  experiments, 
MOrner  never  obtained  higher  results  for  permanganate  used.  Consequently 
the  method  of  washing  described  completely  removes  tartaric  and  thio-acetic 
acids. 

^  "Official  Directions  for  the  Chemical  Examination  of  Wine"  of  June  25th, 
1896.     (German.) 

8  "Agreements  in  regard  to  uniformity  in  inspecting  and  testing  foods,  house- 
hold supplies  and  other  articles  used  in  the  German  Empire  "  Heft  I,  36. 

*  Method  of  Ch.  Girard,  Zeitschrift  fur  analytische  Chemie  22,277  (1SS3)  and 
the  above  "Agreements'"  Heft  I,  62. 


244  DETECTION   OF   POISONS 

cc.  of  ether,  evaporate  the  ether,  dissolve  the  residue  in  5  cc. 
of  hot  water  and  test  the  filtered  solution  for  salicylic  acid  with 
dilute  ferric  chloride  solution  (sp.  gr.  i. 005-1. 010). 

Maltol 

Maltol,  CeHeOs,^  is  formed  in  the  preparation  of  caramel  from  malt,  possibly 
from  maltose  or  isomaltose.  Ether  or  chloroform  extracts  this  substance  from 
the  condensed  vapors  given  off  during  caramelization  and  also  from  beer-wort. 
Maltol  crystallizes  in  monoclinic  prisms  and  plates  from  a  cold  saturated  solution 
in  50  per  cent,  alcohol  (Osann).  Chloroform  gives  denser  crystals.  This  sub- 
stance dissolves  with  difficulty  in  cold  water  or  benzene;  more  readily  in  hot  water, 
alcohol,  ether  or  chloroform;  and  is  insoluble  in  petroleum  ether.  It  dissolves 
in  caustic  alkahne  solutions  but  is  reprecipitated  by  carbon  dioxide.  Maltol 
subhmes  in  shining  leaflets  and  is  volatile  with  water  vapor.  It  reduces  silver 
solution  in  the  cold  and  Fehling's  solution  with  heat.  An  aqueous  maltol  solu- 
tion resembles  saUcyUc  acid  in  becoming  intense  violet  with  ferric  chloride  solu- 
tion, but  differs  from  carbolic  and  salicylic  acids  in  not  turning  red  with  Millon's 
reagent. 

Maltol,  shaken  with  benzoyl  chloride  and  sodium  hydroxide  solution,  gives 
a  mono-benzoyl  derivative  and  consequently  must  contain  one  hydroxy! 
group. 

Aqueous  Chloral  Hydrate  Solution  as  a  Solvent  for  Alkaloids,  Glucosides 

and  Bitter  Principles  and  Its  Use  in  Toxicological  Analysis 

Richard  Mauch 

(Communication  from  Professor  E.  Schaer's  Institute,  Strassburg) 

One  part  of  water  at  17.5°  dissolves  4  parts  of  chloral  hydrate,  forming  a  very 
mobile  solution  which  is  easily  filtered  and  capable  of  being  kept  for  a  long  time 
without  decomposition.  This  80  per  cent,  solution  of  chloral  hydrate  easily 
dissolves  relatively  large  quantities  of  alkaloids  and  glucosides  without  altering 
them  chemically.  At  17.5°  one  part  of  each  of  the  following  substances 
requires  for  solution  the  number  of  parts  of  solvent  stated  in  the  table: 

Chloral  Hydrate 

Solution 

(80%)  Water                 Ether  Chloroform 

Atropine 5  600                         50  3.5 

Quinine 6  2000  freely  soluble  2 

Cocaine 5  700  freely  soluble  freely  soluble 

Morphine 5  5000                     1250  100 

Santonin 4  5000                       125  4 

Strychnine 6.5  6600                     1300  6 

Brucine 6.5  ....                      ....  .... 

Veratrine 7.5  ....                     ....  .... 

^  J.  Brand,  Berichte  der  Deutschen  chemischen  Gesellschaft  27,806  (1894), 
H.  Kiliani  and  M.  Bazlen,  Ibidem  27,  3115  (1894). 


SPECIAL   QUALITATIVE    AND   QUANTITATIVE    METHODS       245 

Caffeine  is  the  only  alkaloid  which  forms  with  chloral  hydrate  a  molecular 
compound  soluble  in  water.  If  a  chloral  hydrate  solution  of  an  alkaloid,  which 
has  been  freshly  prepared  in  the  cold,  is  diluted  with  considerable  water,  the 
unchanged  alkaloid  is  precipitated  almost  quantitatively,  for  instance,  morphine, 
strychnine  and  quinine.  Substances  like  picrotoxin,  santonin  and  acetanilide 
behave  similarly.  But  when  such  solutions  stand  for  a  long  time  at  ordinary 
temperatures,  or  are  heated  for  1-2  hours,  chloral  hydrate  is  decomposed  by 
the  vegetable  base  into  chloroform  and  formic  acid.  Since  the  alkaloidal  salts 
of  formic  acid  are  soluble  in  water,  dilution  with  this  solvent  does  not  precipitate 
the  alkaloids.  R.  Mauch  has  shown  clearly  that  atropine,  brucine,  quinine, 
cocaine,  morphine,  narcotine,  strychnine  and  veratrine  behave  as  just  described. 

In  the  tests  ordinarily  made  with  the  ether  or  chloroform  residue,  R.  Mauch 
recommends  dissolving  the  residue  in  80  or  60  per  cent,  chloral  hydrate  solution. 
The  "chloral  solution"  should  prove  of  great  value  in  color  tests  which  depend 
upon  the  use  of  pure  sulphuric  acid  or  sulphuric  acid  containing  iron  or  molybdic 
acid.  These  solutions  contain  so  little  water  that  it  cannot  modify  the  action 
of  sulphuric  acid  upon  the  substance  in  solution.  Such  a  "chloral  solution"  is 
also  well  adapted  for  zone  tests.  An  aqueous  solution  forms  an  upper  layer  with 
the  "chloral  solution,"  and  the  latter  forms  an  upper  layer  with  concentrated 
sulphuric  acid. 

Specific  gravity  of  80  per  cent,  chloral  hydrate  solution  =  1.514. 
Specific  gravity  of  60  per  cent,  chloral  hydrate  solution  =  1.3535. 

In  the  tests  ordinarily  performed  in  test-tubes,  it  is  best  to  use  small  tube 
(6  or  7  cm.  high;  i  cm.  in  diameter)  holding  6  cc.  They  should  not  be  made  of  too 
thin  glass.  The  chloral  solution  cannot  be  used  in  detecting  picrotoxin,  because 
chloral  hydrate  itself  produces  the  same  reduction  changes  caused  by  picrotoxin. 
The  same  is  true  of  the  test  for  strychnine,  where  sulphuric  acid  and  potassium 
dichromate,  or  any  other  oxidizing  agent,  are  used.  Coniine  and  nicotine  also 
belong  to  the  class  of  alkaloids  which  cannot  be  detected  in  chloral  hydrate  solu- 
tion. Concentrated  chloral  hydrate  solutions  cannot  be  used  directly  in  making 
tests  with  general  alkaloidal  reagents,  because  precipitates  do  not  appear  vmtil 
the  solutions  have  been  diluted  with  6-8  volumes  of  very  dilute  hydrochloric  or 
sulphuric  acid. 

In  using  the  "chloral  hydrate  method"  in  toxicological  analysis,  the  ether, 
chloroform  or  amyl  alcohol  extract  should  be  evaporated  with  gentle  heat  upon  a 
watch  glass  of  medium  size  (about  5  cm.  diameter)  and  not  too  flat.  Add  to  the 
residue,  depending  upon  the  quantity,  about  3  cc.  of  75  per  cent,  chloral  hydrate 
solution.  Cover  the  glass  and  let  it  stand  for  some  time.  Occasionally  tilt 
the  glass  and  bring  the  solution  thoroughly  in  contact  with  the  residue.  Pass  the 
solution  through  a  very  small  filter,  if  necessary,  and  wash  both  watch  glass  and 
filter  with  a  few  drops  of  pure  chloral  hydrate  solution.  Use  this  chloral  hydrate 
solution  for  the  individual  tests.  In  testing  for  strychnine,  evaporate  a  part  oi 
the  chloral  solution  to  dryness  upon  the  water-bath.  Warm,  imtil  the  residue 
does  not  smell  of  chloral,  and  then  test  for  strjxhnine  with  sulphuric  acid  and 
potassium  dichromate. 

To  recover  from  the  chloral  hydrate  solution  most  of  the  alkaloids  and  sub- 
stances hke  picrotoxin,  acetanilide  and  phenacetine,  add  excess  of  sodium  hy- 


246  "        DETECTION   OF   POISONS 

droxide  solution  and  extract  thoroughly  with  a  little  chloroform.  The  "chloral 
hydrate  method"  is  conducive  to  very  neat  work  and  this  is  a  great  advantage. 
The  use  of  metallic  utensils  like  knives  and  spatulas  is  entirely  unnecessary. 

ESTIMATION  OF  ALKALOIDS 

1.  Picrolonate  Method  of  H.  Matthes^ 

Knorr^  gave  the  name  pier  clonic  acid  to  i-p-nitrophenyl-3- 
methyl-4-isonitro-5-pyrazolone.  This  compound  is  formed  by 
the  action  of  nitric  acid  upon  methyl-phenyl-pyrazolone. 
Picrolonic  acid  resembles  picric  acid  iri  its  properties  and  is 
characterized  by  forming  crystalline  salts  with  many  organic 
bases,  as  the  alkaloids.  As  a  rule  these  salts  dissolve  with 
difficulty  and  are  yellow  or  red.  Heat  causes  their  decomposi- 
tion. Picrolonic  acid  is  frequently  of  service  in  characterizing 
bases.  Hydrochloric  acid  precipitates  this  compound  from  a 
solution  of  its  sodium  salt  as  a  yellow,  mealy  powder,  melting 
when  rapidly  heated  at  about  128°,  becoming  dark  in  color  and 
undergoing  decomposition  with  rapid  evolution  of  gas.  Knorr 
first  gave  picrolonic  acid  formula  I  but  now  formula  11^  is 
preferred: 

I.  NO2.C6H4.N  II.  NO2.C6H4.N 

/\  /\ 

N     C.OH  N    CO 

li       II  II      I        ^o 

CH3.C— C.NO2  CH3.C— C=Nf 

H.  Matthes  has  estimated  many  alkaloids  quantitatively  by 
means  of  picrolonic  acid.  Collect  the  precipitated  alkaloidal 
picrolonate  in  a  weighed  Gooch  crucible,  wash,  dry  and  weigh. 
Estimation  of  alkaloids  is  possible  by  this  method,  because  the 
picrolonates  are  constant  in  composition.  Morphine,  hydras- 
tine,  codeine,  strychnine,  brucine,  pilocarpine  and  stypticine^ 
can  be  quantitatively  estimated  by  this  method. 

^  H.  Matthes  and  O.  Rammstedt,  Zeitschrift  fiir  analytische  Chemie  46,  565 
(1907)  and  Archiv  der  Pharmazie  245,  112  (1907). 

2  Berichte  der  Deutschen  chemischen  Gesellschaft  30,  914  (1897). 

2  R.  Zeine,  Inaugural  Dissertation,  Jena,  1906. 

*  Stypticine  =  cotarnine  hydrochloride,  C12H16NO4.HCI.H2O. 


SPECIAL  QUALITATIVE    AND  QUANTITATIVE   METHODS       247 

Estimation  of  Morphine,  Codeine  and  Stypticine  in  Solutions,  Tablets 
and  Sugar  Triturations 

Dissolve  the  weighed  trituration  or  tablet  in  the  smallest 
quantity  of  water  possible  and  add  picrolonic  acid  solution 
(about  O.I  n-solution  in  alcohol)  in  slight  excess.  The  picro- 
lonate  separates  at  once,  or  very  soon,  as  yellow  crystals  or  a 
crystalline  meal.  Cool  15-30  minutes  in  ice  water  and  collect 
the  precipitate  in  a  weighed  Gooch  crucible.  Wash  with  a 
little  ice  water,  dry  30  minutes  at  110°  and  weigh.  Morphine 
picrolonate  usually  separates  in  10-30  minutes.  Cooling  aids 
precipitation. 

Formula  Mol.  Wt.  Decomposition-point 

Morphine  picrolonate :      C17H19NO3.C10H8N4O6.         549         200-210° 
Codeine  picrolonate:         C1SH21NO3.C10H8N4O6.         563         about  225° 
Cotarnine  picrolonate:      C12H18NO4.C10H8N4O6.        501         205-210° 

Notes.' — Practice  Analyses:  Morphine  powder:  0.01-0.02  gram  morphine  hy- 
drochloride, C17H19NO3.HCI.3H2O  +  0.5  gram  sugar.  0.2-0.5  gram  codeine 
phosphate,  C18H21NO3.H3PO4.2H2O  +  0.5  gram  sugar.  Stypticine  tablets  E. 
Merck. 

Do  not  use  too  dilute  solutions  of  the  alkaloids  in  these  determinations  and  do 
not  wash  the  picrolonate  precipitates  with  too  much  water.  Dissolve  the  pow- 
dered morphine  and  sugar  mixture  in  about  5-10  cc.  of  water.  Matthes  and 
Rammstedt  in  examining  the  morphine  powder  obtained  the  following  results: 

Weights  taken:  0.019  morphine  hydrochloride  +  0.5  gram  sugar. 
Results  obtained: 

I.  0.0273  gram  morphine  picrolonate  =  0.0187    gram   morphine  hydro- 
chloride. 

II.  0.0274  gram   morphine    picrolonate  =   0.0187    gram     morphine  hy- 
drochloride. 

In  a  second  experiment  every  10  cc.  of  an  aqueous  solution  contained  0.0104 
gram  of  morphine  hydrochloride. 
Results  obtained: 

I.  o.or47   gram  morphme   picrolonate  =  o.oioi  gram    morphine   hj-dro- 
chloride. 

II.  0.0146  gram  morphine   picrolonate  =  0.0099   gram    morphine  hydro- 
chloride.' 

The  application  of  the  picrolonate  method  to  the  estimation  of  hydrastine  in 
hydrastis  root  and  extract,  of  nux  vomica  alkaloids  in  nux  vomica  and  extract  and 
of  pilocarpine  in  jaborandum  leaves  is  described  in  Chapter  VI  (see  pages  275, 
279  and  289. 

1  Professor  Matthes  has  kindly  stated  that  this  picrolonate  method  gives  less 
satisfactory  results  with  smaller  quantities  of  morphine  (0.005  and  less). 


248  DETECTION   OF  POISONS 

2.  Estimation  of  Alkaloids  by  Means  of  Potassium  Bismuthous  Iodide 

(H.  Thomsi) 

Dissolve  the  particular  alkaloid  in  sulphuric  acid  and  pre- 
cipitate completely  with  potassium  bismuthous  iodide  prepared 
as  described  by  Kraut. ^  Decompose  the  precipitate  with  a 
mixture  of  sodium  carbonate  and  hydroxide,  extract  the  free 
alkaloid  with  ether  and  weigh.  By  this  method  Thoms  has 
recovered  atropine,  hyoscyamine,  scopolamine,  strychnine, 
quinine,  caffeine  and  antipyrine  from  their  potassium  bis- 
muthous iodide  precipitates  unaltered  and  nearly  quantitatively. 
He  has  also  used  this  method  with  success  in  estimating  quanti- 
tatively the  alkaloids  in  belladonna  extract. 

Procedure. — Dissolve  the  alkaloidal  salt,  or  2  grams  of  bella- 
donna extract,  in  50  cc.  of  water.  Add  first  10  cc.  of  10  per  cent, 
sulphuric  acid,  stir  and  precipitate  with  5  cc.  of  potassium  bis- 
muthous iodide  solution.  Collect  the  precipitate  upon  a  dry 
filter  and  wash  twice  with  5  cc.  portions  of  10  per  cent,  sulphuric 
acid.  Transfer  the  thoroughly  drained  precipitate  and  paper 
to  a  wide-mouth  extraction  cylinder  having  a  tight  glass  stopper. 
Add  0.3  gram  of  sodium  sulphite,  then  30  cc.  of  15  per  cent,  so- 
dium hydroxide  solution  and  shake.  Add  quickly  15  grams  of 
sodium  chloride  and  100  cc.  of  ether.  Shake  frequently  and  let 
stand  for  3  hours.  The  ether  contains  the  alkaloid  and  settles 
well.  Remove  with  a  pipette  50  cc.  of  the  ether  solution  (  = 
half  the  solution  of  the  alkaloidal  salt,  or  i  gram  of  belladonna 
extract)  and  titrate  this  ether  solution  in  a  flask  with  o.oi 
n-hydrochloric  acid,  using  iodeosine  as  indicator. 

After  titrating  the  belladonna  alkaloids,  use  in  the  calculation 
the  equivalent  weight  of  atropine-hyoscy amine,  C17H23NO3  = 
289.  1000  cc.  of  0.01  n-hydrochloric  acid  correspond  to  2.89 
grams  of  atropine-hyoscyamine.  Atropine  and  hyoscyamine 
being  isomeric,  monacid  bases,  their  formula  weight  and  equiva- 
lent weight  are  the  same. 

1  Berichte  der  Deutschen  pharmazeutischen  Gesellschaft  13,  240  (1903);  15,  85 
(1905);  16,  130  (1906)  (D.  Jonescu). 

2  Annalen  der  Chemie  und  Pharmazie  210,  310  (1882).  See  "Preparation  of 
Reagents,"  page  311. 


SPECIAL  QUALITATIVE    AND   QUANTITATIVE   METHODS        249 

Quinine,  caffeine  and  antipyrine  were  also  recovered  un- 
altered from  potassium  bismuthous  iodide  precipitates.  After 
decomposition  of  the  precipitates,  they  were  obtained  almost 
quantitatively  but  were  estimated  gravimctrically. 

Dissolve  2  grams  of  quinine^  in  50  cc.  of  water  acidified  with 
sulphuric  acid  and  precipitate  with  potassium  bismuthous  iodide. 
Filter  the  precipitate  with  suction  and  wash  with  5  per  cent, 
sulphuric  acid.  Transfer  precipitate  and  paper  to  an  extraction 
cylinder  and  shake  thoroughly  with  a  mixture  of  20  grams  of 
crystallized  sodium  carbonate  and  40  cc.  of  10  per  cent,  sodium 
hydroxide  solution.  The  yellowish  red  precipitate  gradually 
becomes  white.  Add  30  cc.  of  ether  and  shake  well  for  30 
minutes.  Pipette  off  25  cc.  of  the  clear  ether  solution,  evap- 
orate in  a  weighed  glass  dish,  dry  the  residue  at  100°  and  weigh. 
The  weight  of  quinine  was  0.9405  instead  of  i  gram. 

Caffeine  was  estimated  in  the  same  way,  except  that  this 
alkaloid  was  extracted  with  chloroform  after  decomposition  of 
the  potassium  bismuthous  iodide  precipitate  with  alkahne 
hydroxide  and  carbonate.  The  weight  of  caffeine  was  0.9546 
instead  of  i  gram. 

The  precipitate  obtained  by  adding  potassium  bismuthous 
iodide  to  a  solution  of  antipyrine  in  sulphuric  acid  (10  per  cent. 
H2SO4)  is  not  decomposed  as  easily  as  are  those  of  quinine  and 
caffeine.  The  precipitate  from  2  grams  of  antipyrine  must  be 
shaken  i  hour  with  20  grams  of  sodium  carbonate  and  60  cc. 
of  10  per  cent,  sodium  hydroxide  solution.  Antipyrine  must  be 
extracted  with  chloroform.  The  weight  of  antipyrine  was 
0.9273  instead  of  i  gram. 

Notes. — Potassium  bismuthous  iodide  precipitates  fixed  and  volatile 
alkaloids  but  not  ammonium  salts.  If  the  estimation  of  volatile  bases  is  unnec- 
essary, as  in  the  examination  of  belladonna  extract,  evaporate  the  50  cc.  of  ether 
extract  (see  above)  upon  the  water-bath.  Warm  the  residue  and  in  a  few  min- 
utes the  strong  narcotic  odor  of  volatile  bases  will  disappear.  Dissolve  the 
residue  in  a  little  acid-free  alcohol  and  dilute  with  ether.  Before  using  a  flask 
for  titrations  carefully  test  it  beforehand  for  alkahnity.  If  a  positive  test  is 
obtained,  alkalinity  must  be  removed.  An  odor  like  iodoform,  probably  due  to 
the  action  of  sodium  hypoidite  upon  the  alkaloid,  has  been  observed  when  sodi- 

^  According  to  experiments  of  D.  Jonescu  (Joe.  cit.). 


250  DETECTION   OF   POISONS 

um  hydroxide  solution  acts  upon  potassium  bismuthous  iodide  precipitates. 
Addition  of  sodium  sulphite  may  prevent  this  action.  After  addition  of  sodium 
chloride  ether  takes  up  the  alkaloid  more  readily.  But  vigorous  shaking  is 
always  needed  to  cause  complete  transfer  of  alkaloid  to  the  ether, 

3.  Estimation  of  Alkaloids  by  H.  M.  Gordin^ 

Gordin  has  found  that  periodides  of  the  alkaloids,  whatever  be  their  composi- 
tion, when  precipitated  from  aqueous  solution  by  iodo-potassium  iodide  in  pres- 
ence of  acids,  always  contain  one  equivalent  of  combined  acid  for  every  molecule  of 
monacid  alkaloid.  These  periodides  have=the  general  formula  (Alkaloid,  HI)mIn. 
Iodo-potassium  iodide,  added  to  a  solution  of  a  monacid  alkaloid  acidified  with 
hydrochloric  acid,  first  gives  an  alkaloid  hydrochloride,  changed  by  potassium 
iodide  to  alkaloid  hydriodide  and  finally  precipitated  as  insoluble  periodide  by  tak- 
ing up  iodine: 

(a)  Alkaloid  +  HCl  =  Alkaloid.HCl, 

(/3)   AIka]oid.HCl      +    KI  =  Alkaloid.HI  +  KCl, 

(7)  m(Alkaloid.HI)  -J-     I^  =  (Alkaloid.HI)jj^In  =  Precipitate. 

In  the  precipitation  of  an  alkaloid  in  acid  solution  with  iodo-potassium  iodide, 
one  equivalent  of  acid  goes  with  the  precipitate  and  disappears  from  solution. 
In  many  cases  potassium  mercuric  iodide  may  be  substituted  to  advantage  for 
iodo-potassium  iodide.  Gordin  has  found  that  the  composition  of  the  precipitate 
changes  only  as  regards  mercuric  iodide  and  not  as  far  as  acid  is  concerned,  for  in 
this  case  also  the  precipitate  contains  one  equivalent  of  acid  for  a  monacid  alkaloid. 
Use  in  the  titration  0.05  n-hydrochloric  acid  and  0.05  n-potassium  hydroxide 
solution.  Prepare  a  solution  containing  a  weighed  quantity  of  pure  alkaloid,  for 
example,  pure  morphine.  Dissolve  about  0.2  gram  of  chemically  pure  morphine, 
previously  completely  dehydrated  at  120°,  in  30  cc.  of  0.05  n-hydrochloric  acid 
in  a  100  cc.  volumetric  flask.  Shake  and  add  gradually  iodo-potassium  iodide 
to  this  solution,  until  precipitation  ceases  and  the  supernatant  liquid  is  dark  red. 
Dilute  to  the  100  cc.  mark  with  water  and  shake  vigorously  until  the  liquid  above 
the  precipitate  is  entirely  clear.  Pass  50  cc.  of  solution  through  a  dry  filter,  de- 
colorize the  filtrate  with  a  few  drops  of  sodium  thiosulphate  solution  and  titrate 
excess  of  0.05  n-hydrochloric  acid  with  0.05  n-potassium  hydroxide  solution,  using 
phenolphthalein  as  indicator.  Calculate  from  the  result  how  many  grams  of 
morphine  have  been  neutralized  by  i  cc.  of  the  acid.  Comparison  of  the  equiva- 
lent weight  of  morphine  with  that  of  any  other  monacid  alkaloid  gives  the  corre- 
sponding factor  to  be  used  in  the  calculation.  For  example,  Gordin  found  in  his 
experiments  that  i  cc.  of  approximately  0.05  n-hydrochloric  acid  neutralized 
0.0137  gram  of  anhydrous  morphine,  C17H19NO3.  The  factor  (x)  for  strychnine, 
C21H22N2O2  (=  334),  which  is  also  a  monacid  base,  is  as  follows: 

Morphine  :  Strychnine 

285       '■       334  =  0-OI37   '■  ^      (x  =  0.0160) 

^  Berichte  der  Deutschen  chemischen  Gesellschaft  32,  2871  (1899).  Archiv  der 
Pharmazie  238,  335  (1900).  Gordin  and  A.  B.  Prescott,  Archiv  der  Pharmazie 
237,  380  (1899). 


SPECIAL  QUALITATIVE    AND   QUANTITATIVE    METHODS       251 

and  that  for  the  monacid  base  cocaine,  Ci/'l2iN04(=  303),  according  to   the 
proportion  is: 

Morphine  :  Cocaine 

28s      :    303  =  0.0137  :  X       (x=  0.0146). 

If  potassium  mercuric  iodide  is  used  to  precipitate  an  alkaloid,  the  method  is 
the  same  as  with  iodo-potassium  iodide  except  that  there  is  no  need  of  treating 
the  50  cc.  of  filtered  solution  with  thiosulphate. 

Berberine  and  colchicin  cannot  be  estimated  by  Gordin's  method. 

Quantitative  Estimation  of  Strychnine  and  Quinine  Together 
(E.  F.  Harrison  and  D.  Gair^ 

Occasionally  a  small  amount  of  strychnine  must  be  estimated  in  presence  of  a 
relatively  large  quantity  of  quinine,  as  in  certain  pharmaceutical  preparations.'^ 
Separation  of  the  two  alkaloids  is  possible  by  means  of  Rochelle  salt.  Quinine 
tartrate,  (C2oH24N202)2.C4H60c.2H20,  being  difficultly  soluble  in  water,  forms 
a  white  crystalline  precipitate,  whereas  strychnine  tartrate  remains  in  solution. 

ProcediU'e. — Render  the  solution  of  the  mixed  alkaloids  in  about  40  cc.  of 
water  faintly  acid  with  sulphuric  acid.  Add  enough  ammonia  to  cause  a  slight 
turbidity,  then  15  grams  of  solid  Rochelle  salt  and  more  ammonia,  still  leaving 
the  liquid  acid  to  litmus  paper.  Heat  for  1 5  minutes  upon  the  water-bath  and  then 
set  aside  for  2  hours  until  entirely  cold.  Filter  precipitated  quinine  tartrate  by 
suction  and  wash  with  aqueous  Rochelle  salt  solution  (15  grams  of  salt  in  45  cc. 
of  water)  containing  1-2  drops  of  dilute  sulphuric  acid.  To  determine  strychnine, 
add  sodium  hydroxide  solution  to  the  combined  filtrate  and  wash  water  from 
quinine  tartrate  until  the  reaction  is  alkaline.  Extract  2-3  times  with  chloro- 
form, pour  the  chloroform  extract  through  a  dry  filter  and  distil  in  a  weighed 
flask  to  about  4  cc.  Add  10  cc.  of  absolute  alcohol  and  evaporate  to  drj-ness 
upon  the  water-bath.  To  remove  quinine  still  adhering  to  the  strjxhnine,  ex- 
tract ^the  dry  residue  2-3  times  with  i  cc.  portions  of  ether,^  dry  at  100°  and 
weigh.     This  residue  consists  of  pure,  quinine-free  strychnine. 

Estimation  of  Toxicity  of  Chemical  Compounds  by  Blood  Haemolysis 

(A.  J.  J.  Vandevelde*) 

Vandevelde  originally  used  living  cells  of  a  variety  of  Allium  cepa  (red  Bruns- 
wick onion),  the  cell  membrane  of  which  is  rich  in  anthocj^an.  The  presence  of 
this  substance  obviated  the  necessity  of  using  a  special  coloring  matter  in 
determining  plasmolytically  the  toxicity  of  alcohols,  ethereal  oils  and  other  sub- 
stances.^ Vandevelde  has  recently  recommended  determining  the  toxicity  of 
chemical  compounds  by  blood  hjemolysis,  using  for  this  purpose  defibrinated  ox 

^  Pharmaz.  Journ.  (4)  17,  165. 

^  Compound  Syrup  of  Hypophosphites,  U.  S.  P. 

^  More  ether  dissolves  a  weighable  quantity  of  strjxhnine. 

■*  Chemiker  Zeitung  29,  565  (1905). 

^  Bulletin  de  1' Association  Belgee  de  Chimie  17,  253. 


252  DETECTION   OF   POISONS 

blood  (see  pages  216  and  222).  To  establish  the  toxicity  of  different  alcohols,  the 
concentration  at  which  haemolysis  just  ceases  is  determined.  A  solution,  in  which 
blood  corpuscles  are  not  hydrolyzed  after  a  definite  time  but  are  hydrolyzed  upon 
addition  of  the  slightest  trace  of  the  substance  being  examined,  is  a  non-toxic  solu- 
tion for  blood  corpuscles,  called  by  Vandevelde  a  "critical  solution." 
The  estimation  requires : 

1 .  A  solution  of  0.9  per  cent,  sodium  chloride  in  50  per  cent,  alcohol  by  volume.^ 

2.  An  aqueous  0.9  per  cent,  sodium  chloride  solution. 

3.  A  suspension  of  5  per  cent,  defibrinated  ox  blood  m  0.9  per  cent,  aqueous 
sodium  chloride  solution. 

Experiments  are  made  in  test-tubes.  Place  first  in  each  of  several  tubes  2.5  cc. 
of  the  mixtures  (of  different  concentration)  of  alcoholic  and  aqueous  sodium 
chloride  solution  and  then  2.5  cc.  of  the  suspended  blood.  The  end  point  for 
the  appearance  of  hemolysis  was  set  at  three  hours. 

Vandevelde's  experiments  with  ethyl  alcohol  gave  the  following  results; 

^       r    ,     ■,    ,.  Alcoholic  con- 

Cc.  of  alcoholic 


Cc.  of  sus- 
pended blood 

NaCl  solu- 
tion 

Cc.  of  aqueous 
NaCl  solution 

centration  of 
mixture  in  vol.- 

After  3 
hours 

per  cent. 

2.5 

2 .  20 

0.30 

22.0 

Haemolysis 

2.5 

2.15 

0.35 

21. 5 

Haemolysis 

2.5 

2  .  10 

0.40 

21.0 

Haemolysis 

2-5 

2.05 

0.45 

20.5 

Haemolysis 

2-5 

2  .00 

0.50 

20.0 

Haemolysis 

2-5 

1-95 

o-SS 

19s 

No  haemolysis 

2.5 

1 .90 

0.60 

19.0 

No  haemolysis 

Consequently  the  critical  solution  of  ethyl  alcohol  is  one  containing  19.5  cc.  of 
absolute  alcohol  (C2H6O)  in  100  cc,  or  15.489  grams  of  C2H6O  in  100  cc.  The 
specific  gravity  of  absolute  alcohol  being  0.7943,  19.5  cc.  weigh  19.5  X  0.7943  = 

iS-489- 

According  to  Vandevelde's  experiments  addition  of  methyl  alcohol  diminished 
the  toxicity  of  ethyl  alcohol,  whereas  the  higher  alcohols  were  found  to  be  more 
toxic  than  the  latter.  If  the  toxicity  of  100  parts  by  weight  of  ethyl  alcohol  is 
taken  as  100,  then  47  parts  by  weight  of  isopropyl,  29  parts  of  isobutyl  and  12.5 
parts  of  amyl  alcohol  are  isotoxic  with  that  quantity  of  ethyl  alcohol.  Using 
theplasmolytic  method  with  onion  cells,  Vandevelde  obtained  the  following  results 
with  the  same  series: 

100,  36.8,  21.2,  12.6. 

The  haemolytic  method  is  easily  performed  in  test-tubes  and  does  not  require 
the  use  of  the  microscope.  The  form  of  the  tube,  especially  its  diameter,  is  quite 
important  in  these  experiments.  The  speed  of  haemolysis  increases  with  the 
diameter  of  the  tube.  The  quantity  of  the  blood  corpuscles  is  of  slight  influence 
except  in  narrow  tubes  and  at  the  beginning  of  the  reaction.  Vandevelde  applies 
the  term  "critical  coefi&cient"  to  the  number  giving  the  concentration  of  a  sub- 
stance necessary  to  kiU  the  cells. 

^  The  specific  gravity  of  such  an  alcohol  at  15°  is  0.9348. 


CHAPTER  VI 

QUANTITATIVE    ESTIMATION    OF    ALKALOIDS    AND    OTHER 

PRINCIPLES 

Estimation  of  Alkaloids  in  Drugs  and  Pharmaceutical  Preparations 

(German  Pharmacopoeia) 

Alkaloids  are  nitrogenous  bases  occurring  in  plants.  The 
term  "plant  base"  is  synonymous  with  alkaloid.  The  plant 
families  especially  rich  in  alkaloids  are  berberideas,  cinchonaceae, 
papaveraceas,  solanaceae  and  strychnaceae.  Alkaloids  as  a  rule 
are  not  uniformly  distributed  in  all  parts  of  plants.  They 
occur  most  often  in  roots,  fruits  and  seeds.  If  the  plant  is  a 
tree,  there  is  often  more  alkaloid  in  the  bark  than  in  other  parts. 
The  particular  part  of  the  plant  usually  contains  only  a  few  per 
cent,  of  alkaloid.  Quinine  bark  is  an  exception,  the  quantity 
of  alkaloid  being  5-10  per  cent,  and  sometimes  more.  Plants  as 
a  rule  do  not  contain  free  alkaloids  but  their  salts.  They  are 
combined  not  only  with  the  mineral  acids,  sulphuric,  hydro- 
chloric and  possibly  phosphoric,  but  with  organic  acids,  as  mahc, 
aconitic,  tannic,  citric,  quinic  and  meconic.  Most  free  plant 
bases  dissolve  only  sHghtly  in  water  but  readily  in  ether  and 
chloroform.  The  German  Pharmacopoeia  prescribes  a  mixture 
of  ether  and  chloroform  for  the  extraction  of  free  alkaloids. 
The  finely  powdered  drug  should  first  be  treated  with  a  solution 
of  sodium  hydroxide,  ammonia  or  sodium  carbonate  to  liberate 
alkaloids  from  their  salts: 

C20H24N2O2 
I 
C6H7(OH)4COOH  +  NaOH  =  C20H24N2O2  +  C6H7(OH)4COONa  +  H2O. 

Quinine  quinate^  Quinine  Sodium  quinate 

The  ether-chloroform  mixture  removes  not  only  alkaloids 
from  the  drug  but  varying  amounts  of  other  substances,  as  fat, 
resin,  wax  and  pigments.     To  free  the  alkaloid  from  such  im- 

^  Cinchona  bark  contains  quinine  in  the  form  of  this  salt. 

253 


254  DETECTION    OE    POISONS 

purities,  shake  the  ether-chloroform  extract  with  a  measured 
excess  of  a.i  or  o.oi  n-hydrochloric  acid.  The  alkaloid  passes 
into  aqueous  solution  as  hydrochloride : 

C20H24N2O2  +  HCl  =  C20H24N2O2.HCI 

Quinine  Quinine  hydrochloride 

But  the  impurities  remain  in  the  ether-chloroform  mixture. 
Finally,  determine  excess  of  hydrochloric  acid  by  titration  with 
0.1  or  0.01  n-potassium  hydroxide  solution,  employing  usually 
iodeosine  as  indicator.  Calculate  the  amount  of  alkaloid  in  the 
drug  from  the  difference  between  the  original  quantity  of  acid 
and  the  excess. 

The  estimation  of  alkaloids  in  drugs  and  pharmaceutical 
preparations,  according  to  directions  given  by  the  German 
Pharmacopoeia,  requires  the  following  steps: 

1.  Liberation  of  alkaloids  from  salts  by  means  of  stronger 
bases,  as  potassium  and  sodium  hydroxides,  ammonia  and 
sodium  carbonate. 

2.  Extraction  of  free  alkaloids  with  ether-chloroform  mixture. 

3.  Transference  of  alkaloids  from  ether-chloroform  to  aque- 
ous o.i  or  0.01  n-hydrochloric  acid  solution. 

4.  Determination  of  excess  of  hydrochloric  acid  in  an  ahquot 
volume,  usually  50  cc.  of  the  hydrochloric  acid  solution  of  the 
alkaloid  diluted  to  100  cc,  by  titration  with  o.i  or  o.oi  n-potas- 
sium hydroxide  solution. 

Alkaloids  in  Aconite  Root 

Officinal  aconite  root  is  the  root  of  Aconitum  Napellus  col- 
lected at  the  end  of  flowering.  Two  alkaloids  are  present, 
namely,  aconitine,  C34H47NO11,  and  pier aconi tine,  C31H47NO10 
(?),  characterized  by  its  very  bitter  taste.  Both  alkaloids  are 
combined  with  aconitic  acid, 

CH.COOH 
C.COOH       . 

I 

CHo.COOH 


QUANTITATIVE   ESTIMATION    OF    ALKALOIDS  255 

Boiled  with  water,  or  alcoholic  potassium  hydroxide  solution, 
aconitine  yields  a  new  base,  aconinc,  benzoic  and  acetic  acids  :^ 

Csjr^rNOn  +  2H2O  =  C26H41NO9  +  C2H4O2  +  CeHa.COOH. 

Aconitine  Aconine  Acetic  Benzoic  acid 

acid 

This  reaction  presents  aconitine  as  acetyl-benzoyl-aconine, 

/COCH3 

C26H39N09< 

\COC0H5 

Since  aconitine  has  been  shown  to  contain  four  methoxyl 
groups,  the  formula  of  this  alkaloid  may  be  written: 

/COCH3 

C2lH27(OCH3)4N05< 

^COCeHs 

Aconine,  therefore,  is  C2iH27(OCH3)4(OH)2N03,  and  picra- 
conitine  must  be  regarded  as  benzoyl-aconine,  having  the  for- 
mula C2iH2i(OCH3)4(OH)N04(COC6H5). 


Estimation  of  Aconitine 

(German  Pharmacopoeia) 

Place  12  grams  of  rather  finely  powdered  aconite  root  dried  at  100°  in  an 
Erlenmeyer  flask  and  add  90  grams  of  ether  and  30  grams  of  chloroform.  Shake 
well  and  add  10  cc.  of  a  mixture  of  2  parts  of  sodium  hydroxide  solution  and  i 
part  of  water.  Let  the  mixture  stand  3  hours,  shaking  vigorously  at  frequent 
intervals.  Then  add  10  cc.  of  water,  or  enough  to  cause  the  powder  to  gather 
into  balls  after  vigorous  shaking  and  leave  the  supernatant  ether-chloroform 
solution  perfectly  clear.  After  an  hour  pass  100  grams  of  the  clear  ether- 
chloroform  solution  through  a  dry  filter  kept  well  covered  and  receive  the  filtrate 
in  a  small  flask.  Distil  about  half  the  solvent  and  pour  the  remainder  into  a 
separating  funnel.  Wash  the  flask  with  three  5  cc.  portions  of  a  mixture  of  3 
parts  of  ether  and  i  part  of  chloroform,  and  thoroughly  shake  the  combined 
solutions  with  25  cc.  of  o.oi  n-hydrochloric  acid.  When  the  liquids  have 
separated  perfectly  clear,  add  enough  ether  to  bring  the  ether-chloroform  solu- 
tion to  the  surface.  Pass  the  acid  solution  through  a  small  filter  moistened  -^-ith 
water  and  collect  the  filtrate  in  a  100  cc.  flask.  Make  three  more  extractions 
of  the  ether-chloroform  solution  ^^•ith  10  cc.  portions  of  water,  and  pass  these 
extracts  through  the  same  filter.  Wash  the  latter  with  water  and  dilute  the  total 
solution  with  water  to  100  cc.  Place  50  cc.  of  tliis  solution  in  a  200  cc.  flask  with 
50  cc.  of  water  and  enough  ether  to  make  a  layer  about  i  cm.  thick.  Add  5 
drops  of  iodeosine  solution  and  run  in,  while  shaking,  enough  o.oi  n-potassium 
hydroxide  solution  to  turn  the  aqueous  solution  pale  red. 

^  Freund  and   Beck,   Berichte   der   Deutschen   chemischen   Gesellschaft   27, 

433.  720  (1894);  28,  192,  2537  (1895). 


256  DETECTION   OF  POISONS 

Calculation. — Dissolve  the  aconite  alkaloids  set  free  from  their  salts  by 
sodium  hydroxide  solution  in  120  grams  of  ether-chloroform.  Weigh  100  grams 
of  this  solution  (=  alkaloids  from  10  grams  of  aconite  root).  Dissolve  the 
alkaloids  with  25  cc.  of  o.oi  n-hydrochloric  acid,  bringing  the  volume  to  100  cc. 
Determine  excess  of  acid  in  50  cc.  of  this  solution  (=  alkaloids  from  5  grams  of 
root).  If,  for  example,  this  requires  8.5  cc.  of  o.oi  n-potassium  hydroxide 
solution,  then  the  alkaloids  have  combined  with  12.5  —  8.5  =  4  cc.  of  o.oi  n-acid. 
Since  the  equivalent  weight  of  aconitine  C34H47NO11  =  645,  100  cc.  of  o.oi 
n-hydrochloric  acid  unite  with  6.45  grams  of  aconitine.     The  proportion 

1000  :  6.45  =  4  :  X         (x  =  0.0258) 

shows  that  5  grams  of  aconite  root  contain  0.0258  gram  of  alkaloids,  correspond- 
ing to  0.51  per  cent.  The  German  Pharmacopoeia  demands  this  quantity  of 
aconitine  in  aconite  root  as  a  minimum.  Using  a  different  method,  the  United 
States  Pharmacopoeia  has  the  same  limit. 


Estimation  of  Cantharidin  in  Spanish  Flies 

(German  Pharmacopoeia) 

Place  25  grams  of  Spanish  flies  ground  mediumly  fine  in  an  Erlenmeyer  flask 
and  add  100  grams  of  chloroform  and  2  cc.  of  hydrochloric  acid.^  Shake  the 
mixture  frequently  during  24  hours.  Then  pour  5  2  grams  of  the  chloroform  solu- 
tion through  a  dry  filter  kept  well  covered,  and  collect  the  filtrate  in  a  weighed 
flask  holding  80-100  cc.  Distil  the  chloroform,  and  add  5  cc.  of  petroleum  ether 
to  the  residue.  Stopper  the  flask,  and  let  the  mixture  stand  12  hours,  with  oc- 
casional agitation.  Dry  at  100°  and  weigh  a  filter  (5  cm.  in  diameter).  Pass 
the  liquid  through  this  filter,  having  first  moistened  it  with  petroleum  ether. 
Treat  the  undissolved  residue  twice  with  petroleum  ether,  each  time  using  10  cc. 
and  shaking.  Pass  this  solvent  through  the  same  filter,  and  disregard  crystals 
adhering  to  the  side  of  the  flask.  Dry  the  filter  and  flask,  and  wash  both  with 
a  little  water,  containing  a  drop  of  ammonium  carbonate  solution  to  every  10  cc, 
until  this  solvent  is  only  faintly  yellow.  Finally,  wash  once  with  5  cc.  of  water, 
and  dry  both  flask  and  filter.  Place  filter  and  contents  in  the  flask,  and  dry  at 
100°  to  constant  weight.     The  crystalline  residue  should  weigh  at  least  o.i  gram. 

Notes. — Additional  information  about  cantharidin  is  given 
on  page  196.  Spanish  flies  contain  cantharidin  partly  free  and 
partly  as  an  .alkali  salt  of  cantharidic  acid  (cantharidate) . 
Hydrochloric  acid  sets  cantharidic  acid  free  and  the  latter  then 
passes  at  once  into  cantharidin,  its  internal  anhydride  (lactone). 
Consequently  hydrochloric  acid  is  essential  to  the  determination 
of  that  cantharidin  present  in  Spanish  flies  as  cantharidate. 
Chloroform  not  only  dissolves  cantharidin  but  fatty  substances 

^  Specific  gravity  1.124  =  25  per  cent.  HCl. 


QUANTITATIVE    ESTIMATION    OF   ALKALOIDS  257 

in  the  flies.  To  isolate  pure  cantharidin  from  these  impurities, 
distil  the  chloroform  and  let  the  residue  stand  for  12  hours  in  the 
cold  with  petroletiln  benzene.  Fat  readily  dissolves  but  can- 
tharidin is  as  good  as  insoluble  in  this  solvent.  The  German 
Pharmacopoeia  finally  directs  weighing  the  cantharidin  from 
12.5  grams  of  powdered  Spanish  flies.  The  quantity  should  be 
at  least  o.i  gram,  corresponding  to  0.8  per  cent,  of  cantharidin 
as  a  minimum.  With  sufhcienit  care,  white  crystalline  can- 
tharidin may  be  isolated  from  Spanish  flies. 

Baudin  obtained  from  good  flies  1.06  per  cent,  of  cantharidin, 
of  which  0.72  per  cent,  was  free  and  0.34  per  cent,  combined 
as  cantharidate.  Dieterich  found  only  0.3  per  cent,  of  free 
cantharidin. 

Estimation  of  Cinchona  Alkaloids 

(German  Pharmacopoeia) 

I.  In  Cinchona  Bark. — To  determine  total  alkaloids,  pour  90  grams  of  ether 
and  30  grams  of  chloroform  upon  12  grams  of  finely  ground  cinchona  bark, 
dried  at  100°  and  placed  in  an  Erlenmeyer  flask.  Add  10  cc.  of  sodium  hydroxide 
solution.  Shake  vigorously  at  frequent  intervals  during  3  hours.  Then  add  10 
cc.  of  water,  or  enough  to  cause  the  powdered  cinchona  to  gather  into  lumps 
after  vigorous  shaking,  thus  leaving  the  supernatant  ether-chloroform  solution 
perfectly  clear.  Let  the  ether-chloroform  solution  stand  an  hour,  and  then  pass 
too  grams  through  a  dry  filter,  kept  well  covered.  Collect  the  filtrate  in  a  flask, 
and  distil  half  the  solvent.  Pour  the  remaining  ether-chloroform  solution  into 
a  separating  funnel,  and  wash  the  flask  three  times  with  5  cc.  portions  of  a 
mixture  of  3  parts  of  ether  and  i  part  of  chloroform.  Thoroughly  extract  the 
total  ether-chloroform  solution  with  25  cc.  of  o.i  n-hydrochloric  acid.  When  the 
contents  of  the  separating  funnel  are  perfectly  clear,  add  enough  ether  to  bring 
the  ether-chloroform  solution  to  the  surface.  Pass  the  acid  solution  through  a 
small  filter  moistened  with  water,  and  receive  the  filtrate  in  a  100  cc.  flask. 
Make  three  more  extractions  of  the  ether-chloroform  solution  with  10  cc.  portions 
of  water,  and  pass  these  extracts  through  the  same  filter.  Wash  the  filter  with 
water,  and  bring  the  volume  of  the  filtrate  to  100  cc.  Finally,  measure  50  cc. 
of  this  solution  with  a  pipette,  and  add  freshly  prepared  haematoxylin  solution, 
made  by  dissolving  a  small  particle  of  this  substance  in  i  cc.  of  alcohol.  Shake 
and  add  enough  0.1  n-potassium  hj'droxide  solution  to  give  the  niLxture  a  j-eUowish 
color,  which  quicklj^  changes  after  vigorous  agitation  to  bluish  violet.^ 

Notes  and  Calculation. — Both  quinine  and  quinidine  have 
the  formula  C20H24N2O2  and  cinchonine  and  cinchonidine  the 

^  The  German  Pharmacopoeia  prescribes  that  not   more   than  4.3  cc.  of  0.1 
n-potassium  hydroxide  should  be  required. 
17 


258  DETECTION   OF  POISONS 

formula  C19H22N2O.  These  are  the  most  important  alkaloids 
in  cinchona  bark.  They  are  present  in  all  true  cinchona  barks 
as  salts  of  quinic  acid,  C17H12O6,  and  quino-tannic  acid.  Fuller 
information  regarding  the  chemistry  of  quinine  and  cinchonine 
is  given  on  page  114. 

Quinic  acid  is  widespread  in  the  vegetable  kingdom.  This 
monobasic,  pentatomic  acid,  having  the  formula,  C6H7(OH)4- 
COOH,  is  a  hexahydro-tetroxy-benzoic  acid.  It  crystallizes 
in  large  monocKnic  prisms  melting  at  162°.  As  far  as  the 
chemical  behavior  of  quinic  acid  is  concerned,  either  of  the 
following  formulas  is  possible : 

I.     H     OH  II. 

\/ 
C 

/^ 
H2C      CH.OH 

I        i 
H2C      CH.OH 

\/ 
C 

/\ 
HO      COOH 

The  formation  of  tetra-acetyl-quinic  acid,  (CH3COO)4- 
CgHtCOOH,  and  tetra-benzoyl-quinic  acid,  (C6H5COO)4C6H7- 
COOH,  shows  that  quinic  acid  contains  four  alcoholic  hydroxy! 
groups.  Addition  of  sodium  hydroxide  solution  to  cinchona 
powder  sets  the  alkaloids  free  from  their  salts: 

C20H24N2O2 

C6H7(OH)4COOH  +  NaOH  =  C20H24N2O4  +  H2O  +  C6H7(pH)4CpONa. 

Quinine  quinate  Quinine  Sodium  quinate 

Only  100  grams  of  the  original  120  grams  of  ether-chloroform 
mixture  (  =  12  grams  of  cinchona  powder)  are  in  the  filtrate. 
This  solution  contains  the  alkaloids  in  10  grams  of  bark. 
These  100  grams  are  extracted  with  25  cc.  of  o.i  n-hydrochloric 
acid,  the  alkaloids  passing  into  aqueous  solution  as  hydro- 
chlorides, and  the  volume  is  brought  to  100  cc.  Finally, 
excess  of  o.i  n-hydrochloric  acid  in  50  cc.  (  =  alkaloids  in  5 
grams  of  bark)  of  this  hydrochloric  acid  solution  is  determined 
by  titration.     In  these  determinations  with  very  dilute  hydro- 


H      OH 

\/ 
C 

H2C      CH.OH 

HO.HC      CH2 

C 

HO      COOH 

QUANTITATIVE    ESTIMATION    OF    ALKALOIDS  2o9 

chloric  acid,  cinchona  alkaloids  behave  as  monacid  bases, ^ 
quinine  forming  C2oH24N202-HCl  and  cinchonine  C]9H22N20.- 
HCl. 

The  mean  of  the  equivalent  weight  of  quinine  (324)  and 
cinchonine  (294),  that  is  to  say,  (324  +  294)  divided  by  2  =309, 
may  be  taken  as  the  equivalent  weight.  This  value  agrees 
approximately  with  the  actual  quantities  of  these  alkaloids 
in  cinchona  bark.  Consequently  1000  cc.  of  o.i  n-hydrochloric 
acid  are  equivalent  to  30.9  grams  of  cinchona  alkaloids. 

Example. — Titration  of  50  cc.  of  the  hydrochloric  acid  solution  of  alkaloids,  in 
preparing  which  12.5  cc.  of  o.i  n-hydrochloric  acid  were  used,  required  2.6  cc. 
of  O.I  n-potassium  hydroxide  solution,  equivalent  to  the  volume  of  0.1 
n-hydrochloric  acid  in  excess.  12.5  —  2.6  =  9.9  cc.  of  0.1  n-hydrochloric  acid 
have  combined  with  the  alkaloids  in  5  grams  of  cinchona  bark.     The  proportion 

Cc.  O.I  n-HCl:Grams  of  Alkaloids 

1000         :  30.9  =  9.9 :  X     (x  =  0.30591) 

shows  that  5  grams  of  bark  contain  0.30591  gram  of  alkaloids.  Consequently 
100  grams  of  bark  contain  20  X  0.30591  =6.11  grams  of  alkaloids. 

Titrate  the  filtered  ether-chloroform  solution  of  cinchona  alkaloids  at  once. 
The  solution  should  not  be  exposed  for  any  length  of  time  to  direct  sunlight. 
Otherwise  chloroform  may  give  free  hydrochloric  acid 

CHCI3  +  O  =  COCI2  +  HCl 

which  will  neutralize  alkaloids.  The  decomposition  of  0.05  gram  of  chloro- 
form would  give  enough  hydrochloric  acid  to  neutraUze  0.25  gram  of  cinchona 
alkaloids.  Panchaud^  has  shown  that  such  chloroform  solutions  of  cinchona 
alkaloids  after  standing  1 2  hours  yield  only  80  per  cent,  of  the  total  quantity  of 
alkaloids  originally  present. 

Haematoxylin,  C16H14O6.3H2O,  occurs  in  logwood,  the  heart-wood  of  Hasma- 
toxylon  campechianum.  It  usually  crystallizes  in  colorless,  shining,  quadratic 
prisms  containing  3  molecules  of  water,  more  rarely  in  rhombic  crystals  with  i 
molecule  of  water.  It  dissolves  only  slightly  in  cold  water  but  freely  in  boiling 
water,  alcohol  or  ether.  In  contact  with  air  haematoxj'lin  gradually  becomes 
reddish. 

2.  In  Aqueous  and  Alcoholic  Cinchona  Extracts. — To  determine  total  alkaloids 
in  these  preparations,  dissolve  2  grams  of  the  given  extract  in  an  Erlenmeyer 
flask,  using  5  grams  of  water  and  5  grams  of  absolute  alcohol.     Add  50  grams  of 

^  Quinine  dihydrochloride,  C20H24N2O2.2HCI,  is  formed  bj-  passing  gaseous 
hydrogen  chloride  over  quinine  and  also  by  dissolving  the  monohydrochloride, 
C20H24N2O2.HCI,  in  strong  hydrochloric  acid  with  gentle  heat.  An  aqueous 
solution  of  the  dihydrochloride  has  an  acid  reaction. 

"^  Schweizer  Wochenschrift  fiir  Pharmazie  44,  580. 


260  DETECTION   OF  POISONS 

ether  and  20  grams  of  chloroform,  and,  after  vigorous  shaking,  10  cc.  of  sodium 
carbonate  solution  (i  :3).  Shake  frequently,  and  let  the  mixture  stand  an  hour. 
Then  pass  50  grams  of  the  ether-chloroform  solution  through  a  dry  filter,  kept 
well  covered.  Receive  the  filtrate  in  a  flask,  and  distil  half  the  solvent.  Pour 
the  remainder  into  a  separating  funnel,  wash  the  flask  three  times  with  5  cc. 
portions  of  a  mixture  of  3  parts  of  ether  and  i  part  of  chloroform,  and  then  shake 
the  total  ether-chloroform  solution  with  10  cc.  of  o.i  n-hydrochloric  acid.  When 
the  contents  of  the  separating  funnel  are  perfectly  clear,  add  enough  ether  to  bring 
the  ether-chloroform  solution  to  the  surface.  Pass  the  acid  solution  through  a 
small  filter  moistened  with  water,  and  receive  the  filtrate  in  a  100  cc.  flask.  Make 
three  more  extractions  of  the  ether-chloroform  solution  with  10  cc.  portions  of 
water,  and  pass  these  extracts  through  the  same  filter.  Wash  the  filter  with  water 
and  bring  the  volume  of  the  filtrate  to  100  cc.  Finally,  measure  50  cc.  of  this 
solution  with  a  pipette,  and  add  freshly  prepared  haematoxylin  solution,  made 
by  dissolving  a  small  particle  of  this  substance  in  i  cc.  of  alcohol.  Shake  and  add 
enough  o.i  n-potassium  hydroxide  solution  to  give  the  mixture  a  yellowish  color, 
which  quickly  changes  upon  vigorous  agitation  to  bluish  violet. 

Notes  and  Calculation. — The  alkaloids  in  the  two  cinchona 
extracts  are  set  free  from  their  salts  by  sodium  carbonate: 

C20H24N2O2 
2   I  -f-  Na2C03  =  2C20H24N2O2  +2C6H7(OH)4COONa  +  H2O  +  CO2. 

C6H7(OH)4COOH 

Quinine  Quinine  Sodium 

quinate        _  quinate 

The  alkaloids  from  2  grams  of  extract  are  dissolved  in  75 
grams  of  alcohol-ether-chloroform  mixture.  Two-thirds  of  this 
solution,  or  50  grams.  (=  alkaloids  in  1.33  grams  of  extract) 
are  used  in  the  determination.  The  free  alkaloids  in  this  por- 
tion pass  into  aqueous  solution  as  hydrochloride,  C20H24N2O2. 
HCl,  upon  extraction  with  10  cc.  of  0,1  n-hydrochloric 
acid.  The  excess  of  hydrochloric  acid  in  half  of  this  solution 
diluted  to  100  cc,  that  is  to  say,  in  50  cc.  (  =  alkaloids  in  0.666 
gram  of  extract),  is  finally  determined  by  titration.  If  3.7  cc. 
of  O.I  n-potassium  hydroxide  solution  are  required,  5  —  3.7  = 
1.3  cc.  of  O.I  n-hydrochloric  acid  have  combined  with  the 
alkaloids  in  0.666  gram  of  cinchona  extract.  The  mean 
equivalent  weight  of  quinine  and  cinchonine  (  =  309),  used  in 
the  proportion: 

1000  :30.9  =  1.3  :x     (x  =  0.04017) 

shows  that  1.3  cc.  of  0.1  n-hydrochloric  acid  correspond  to 
0.04017  gram  of  alkaloids,   or  6.03   per  cent.     The  German 


QUANTITATIVE   ESTIMATION    OF   ALKALOIDS  2G1 

Pharmacopoeia  demands  this  quantity  as  a  minimum  for  the 
aqueous  extract  of  cinchona  bark. 

In  the  determination  of  alkaloids  in  alcoholic  cinchona  ex- 
tract, titration  of  excess  of  o.i  n-hydrochloric  acid  in  50  cc. 
should  not  require  more  than  2.3  cc.  of  o.i  n-potassium  hy- 
droxide solution.  Then  5  —  2.3  =  2.7  cc.  of  0.1  n-hydrochloric 
acid  represent  the  alkaloids  in  this  solution.  This  extract  at 
the  minimum  must  contain  12.55  pcr  cent,  of  alkaloids. 

Sulphate  Method  of  Estimating  Quinine  in  Mixtures  of  Cinchona  Alkaloids 

(J.  Carles)! 

This  method  is  especially  recommended  for  practical  purposes 
because  of  its  accuracy  and  simplicity.  Differences  in  the 
solubilities  of  the  sulphates  of  cinchona  alkaloids  in  ammonium 
sulphate  solutions  form  the  basis  of  the  method.  E.  Schmidt 
has  found  that  these  sulphates  have  the  following  solubilities 
in  water  at  15°: 

Quinine  sulphate  1:800         Cinchonine  sulphate  1:65 
Quinidine  sulphate  1:100     Cinchonidine  sulphate  1:97. 

Guareschi  has  found  quinine  sulphate  practically  insoluble 
in  an  ammonium  sulphate  solution,  a  result  which  Hille-  has 
confirmed.  An  addition  of  0.0078  gram  to  the  quantity  of 
quinine  sulphate  obtained  is  necessary  on  account  of  the 
quinine  sulphate  in  the  20  cc.  of  wash  water  used. 

I.  Cinchona  Bark. — Place  1 2  grams  of  finely  powdered  cinchona  bark  dried  at 
100°  in  an  Erlenmeyer  flask  and  add  90  grams  of  ether  and  30  grams  of  chloro- 
form. Add  10  cc.  of  sodium  hydroxide  solution.  Shake  vigorously  at  frequent 
intervals  for  3  hours.  Then  add  10  cc.  of  water,  or  enough  to  cause  the  powdered 
cinchona  to  gather  into  balls  after  thorough  shaking  and  leave  the  supernatant 
ether-chloroform  layer  perfectly  clear.  After  i  hour  pass  100  grams  of  the 
ether-chloroform  solution  through  a  dry  filter  kept  well  covered.  Collect  the 
filtrate  in  a  dry  weighed  flask,  distil  the  ether-chloroform  and  dry  the  flask  at  110° 
to  constant  weight.  The  increase  in  the  weight  of  the  flask  corresponds  to  the 
total  alkaloids  in  10  grams  of  bark. 

Warm  the  alkaloidal  residue  in  the  flask  with  water  and  dilute  sulphuric  acid 

1  Zeitschrift  fiir  analytische  Chemie  9,  467  (1870). 

2  W.  Hille  (ArcMv  der  Pharmazie  241,  54  (1903))  has  reviewed  critically  the 
various  methods  that  have  appeared  thus  far  for  the  estimation  of  quinine  in  pres- 
ence of  other  cinchona  alkaloids. 


262  DETECTION   OF   POISONS 

and  filter  the  solution.  Wash  the  flask  3  times  with  water  containing  sulphuric 
acid  and  pour  the  wash  water  through  the  same  filter.  Dilute  the  filtrate  to  about 
50  CO.,  heat  to' boiling  and  exactly  neutralize  with  ammonia.  Cool  and  after  6 
hours  collect  upon  a  weighed  filter,  or  better  in  a  Gooch  crucible,  the  flocculent 
precipitate  of  quinine  sulphate,  wash  with  20  cc.  of  cold  water,  dry  at  110°  and 
weigh. 

Add  0.0078  gram  to  the  weight  of  quinine  sulphate  found  and  calculate  the 
quantity  of  quinine  in  10  grams  of  cinchona  bark  as  follows: 

C20H24N2O2.H2SO4 :  C20H24N2O2  =  Quinine  sulphate  +  0.0078  :  x. 
(746)  (648)  found 

2.  Cinchona  Extract. — Dissolve  3  grams  of  aqueous  cinchona  extract  in  5  grams 
each  of  water  and  absolute  alcohol  and  place  in  a  measuring  cylinder.  Add  50  cc. 
of  ether,  10  cc.  of  chloroform  and,  after  shaking  vigorously  10  cc.  of  sodium  car- 
bonate solution  (1:3).  Shake  at  frequent  intervals  during  3  hours.  When  two 
layers  have  formed,  bring  the  ether  layer  to  the  75  cc.  mark  with  more  ether. 
Rotate  the  container  carefully  and  evaporate  50  cc.  of  the  clear  ether-chloroform 
layer  in  a  dry  weighed  flask.  Dry  i  hour  at  105°  and  weigh  when  cold.  The  in- 
crease in  weight  corresponds  to  the  total  alkaloids  in  2  grams  of  cinchona  extract. 
There  should  be  at  least  0.12  gram,  or  6  per  cent,  of  alkaloid. 

To  determine  quinine,  pour  very  dilute  sulphuric  acid  over  the  weighed  alka- 
loidal  residue  in  the  flask,  warm  and  filter.  Rinse  the  flask  several  times  with 
very  dilute  sulphuric  acid,  bring  the  volume  to  about  50  cc.  with  water  and  pro- 
ceed as  directed  above  under  cinchona  bark.  Collect  the  quinine  sulphate  in  2 
hours  upon  a  weighed  filter,  or  Gooch  crucible.  The  calculation  is  the  same  as 
for  cinchona  bark. 

Estimation  of  Colchicin  in  Colchiciun  Seed  and  Conns 

(J.  Katz  and  G.  Bredemann^) 

Exhaust  colchicum  seed  or  corms  with  60  per  cent,  alcohol 
and  evaporate  50  grams  of  this  extract  to  20  cc.  Add  0.5 
gram  of  solid  paraffine  and  20  cc.  of  water.  Warm  until  the 
parafhne  is  melted  and  the  alcohol  has  been  completely  expelled. 
Cool  the  liquid  evaporated  to  10-15  cc.  and  pass  through  a 
moist  filter.  Melt  the  paraffine  cake  upon  the  water-bath  with 
10  cc.  of  10  per  cent,  acetic  acid  and  pour  the  cold  Hquid  through 
the  same  filter.  Wash  the  latter,  the  paraffine  cake  and  the 
dish  with  water.  Saturate  the  total  filtrate  with  sodium 
chloride  and  extract  first  with  20  cc.  of  chloroform  and  then  with 
10  cc.  portions  until  a  few  drops  of  the  aqueous  liquid  show 
scarcely  any  turbidity  with  0.05  n-iodine  solution.  Pass  the 
chloroform  solution  through  a  filter  moistened  with  this  solvent 

'  Pharmazeutsche  Zentral-Halle  42,  289  and  Apotheker-Zeitung  18,  817. 


QUANIITATIVE   ESTIMATION   OF    ALKALOIDS 


203 


and  evaporate.  To  expel  chloroform  retained  by  the  colchicin, 
dissolve  the  residue  in  a  little  water  and  filter.  Evaporate  the 
solution  in  a  weighed  dish  and  dry  the  residue  over  sulphuric 
acid  to  constant  weight. 

Note. — Using  this  method,  Bredcmann  obtained  the  following  quantities  of 
colchicin: 


In  seed 
In  corms 
In  fresh  flowers 
In  dry  flowers 


0.46  -0.13  per  cent. 
0.032-0.06  per  cent. 
0.6  per  cent. 

1.8  percent. 


Alkaloids  in  Pomegranate  Bark 

Pomegranate  bark,  the  bark  of  Punica  Granatum,  contains 
the  following  four  alkaloids: 


Pelletierine,  CgHuNO, 
Isopelletierine,  CgHisNO, 


Methyl-pelletierine,  C9H17NO, 
Pseudo-pelletierine,  C9H15NO. 


According  to  Piccini  there  is  still  another  alkaloid  in  the  bark  of 
pomegranate  root  isomeric  with  methyl-pelletierine  and  there- 
fore called  isomethyl-pelletierine. 

Ciamician  and  Silber  have  determined  the  structure  of 
pseudo-pelletierine  which  they  call  n-methyl-granatonine. 
Pseudo-pelletierine  (I)  is  a  ketone  which,  upon  treatment  with 
sodium  amalgam  or  with  sodium  and  alcohol,  adds  two  atoms 
of  hydrogen  and  passes  into  the  corresponding  secondary 
alcohol,  n-methyl-granatoline  (II).  Chromic  and  sulphuric 
acids  oxidize  the  latter  to  n-methyl-granatic  acid  (III) .  Nitro- 
gen can  be  eliminated  by  exhaustive  methylation  and  the  final 
product  is  normal  suberic  acid  (IV) : 

I.  H2C— CH— CH2  II.  H2C— CH— CHo 


+  2H 


H2C    N.CH3CO 
HoC— CH— CH2 

Pseudo-pelletierine  = 
n-methyl-granatonine 

III.  HoC— CH— COOH 
H2C     N.CH3 


H2C— CH— CH2.COOH 

n-methyl-granatic  acid 


H2C      N.CH3CH.OH 

I         I  I  — 

H2C— CH  — CHo 

n-methyl-granatolin 

IV.  HoC— CHo. COOH 

I 
H2C 

I 
HoC— CHo.CHo.COOH 

Normal  suberic  acid 


264  DETECTION   OP  POISONS 

Estimation  of  Alkaloid  in  Pomegranate  Bark 

(German  Pharmacopoeia) 

To  determine  total  alkaloids,  pour  90  grams  of  ether  and  30  grams  of  chloro- 
form upon  12  grams  of  rather  finely  ground  pomegranate  bark,  dried  at  100°  and 
placed  in  an  Erlenmeyer  flask.     Shake  vigorously  and  add  10  cc.  of  a  mixture  of 

2  parts  of  sodium  hydroxide  solution  and  i  part  of  water.    Let  the  mixture  stand 

3  hours,  shaking  vigorously  at  frequent  intervals.  Then  add  10  cc.  of  water, 
which  will  cause  the  powder  to  gather  into  balls  after  vigorous  shaking,  and  leave 
the  supernatant  ether-chloroform  solution  perfectly  clear.  After  an  hour,  pass 
100  grams  of  the  clear  ether-chloroform  solution  through  a  dry  filter,  kept  well 
covered  and  receive  the  filtrate  in  a  separating  funnel.  Extract  this  filtrate 
with  50  cc.  of  o.oi  n-hydrochloric  acid  and  pass  this  acid  solution,  when  perfectly 
clear,  through  a  small  filter  moistened  with  water  into  a  100  cc.  flask.  Make 
three  more  extractions  with  10  cc.  portions  of  water,  and  pass  these  extracts 
through  the  same  filter.  Wash  the  filter  with  water  and  dilute  the  total  solution 
with  water  to  100  cc.  Place  50  cc.  of  this  solution  in  a  200  cc.  flask  with  50  cc. 
of  water  and  enough  ether  to  make  a  layer  about  i  cm.  thick.  Add  5  drops  of 
iodeosine  solution  and  enough  o.oi  n-potassium  hydroxide  solution,  shaking 
vigorously  after  each  addition,  to  give  a  pale  red  color  to  the  lower  aqueous 
solution. 

Calculation. — The  100  grams  of  filtered  ether-chloroform  solution  correspond 
to  ID  grams  of  bark.  The  alkaloids  are  transferred  from  this  ether-chloroform 
solution  to  50  cc.  of  o.oi  n-hydrochloric  acid  which  are  diluted  with  water  to 
100  cc.  The  excess  of  hydrochloric  acid  in  50  cc.  of  this  solution  (  =  alkaloids 
from  5  grams  of  bark)  is  determined  by  titration.  If,  for  example,  11  cc.  of  o.oi 
n-potassium  hydroxide  solution  are  used,  then  25  —  11  =  14  cc.  of  o.oi  n-hydro- 
chloric acid  have  combined  with  the  alkaloids  in  50  cc.  of  the  solution.  If  the 
mean  of  the  equivalent  weights  of  pelletierine  (141)  and  pseudo-peUetierine  (153), 
or  147,  is  used  in  the  calculation,  1000  cc.  of  o.oi  n-hydrochloric  acid  neutralize 
1.47  grams  of  the  mixed  alkaloids.     According  to  the  proportion 

1000  :  1.47  =  14  :  X     (x  =  0.02058) 

5  grams  of  pomegranate  bark  contain  0.02058  gram  of  alkaloids  which  corre- 
sponds to  an  alkaloid  content  of  20  X  0.02058,  or  0.41  per  cent.  The  German 
Pharmacopoeia  demands  this  quantity  of  alkaloids  in  pomegranate  bark  as  a 
minimum. 

Estimation  of  Caffeine  in  Coffee,  Tea,  Cola  Nuts  and  Guarana 

(Literature) 

A.  Hilger  and  A.  Juckenack. — Zur  Bestimmung  des  Kaffeins  in  Kaffee  uiid 
Tee.  Forschungsberichte  iiber  Lebensmittel  und  ihre  Beziehungen  zur  Hygiene 
4,  49-50;  C^  1897  I,  775  and  also  4,  145-154  and  C  1897  II,  233. 

H.  Trillich  and  H.  Gockel. — Beitrage  zur  Kenntniss  des  Kafi^ees  und  der 
Kaffeesurrogate.  Forschungsberichte  iiber  Lebensmittel  und  ihre  Beziehungen 
zur  Hygiene  4,  78-88  and  C  1897  I,  1248. 

^  C  =  Chemisches  Zentralblatt. 


QUANTITATIVE  ESTIMATION   OF   ALKALOIDS  2G5 

L.  Graf. — Ueber  Zusammenhang  von  Kaffelngehalt  und  Qualitat  bei  chines- 
ischen  Tee.  Forschungsberichte  liber  Lebensmittel  und  ihre  Beziehungcn  zur 
Hygiene  4,  88-89,  and  C  1897  I,  1249. 

A.  Forster  and  R.  Riechelmann. — Zur  Bestimmung  des  KafTeins  im  Kaflee. 
Zeitschrift  fiir  offentliche  Chcmic  3,  1 29-131  and  C  1897  I,  1259. 

C.  C.  Keller. — Die  Bestimmung  des  Kaffeins  im  Tee.  Berichte  der  Deutschen 
pharmaceutischen  Gesellschaft  7,  105-112  (1897)  and  C  1897  I,  1134. 

A.  Forster  and  A  Riechelmann. — Zur  Bestimmung  des  Kaffeins  im  Kaffee. 
(Entgegnung.)  Zeitschrift  fur  offentliche  Chemie  3,  235-236  and  C  1897  II, 
436. 

E.  Tassily. — Ueber  ein  neues  Verfahren  zur  Bestimmung  des  KafTeins  im  Kaffee. 
Bulletin  de  la  Soci6t6  chimique,  Paris,  (3)  17,  766-768  and  C  1897  II,  644. 

K.  Dieterich. — Ueber  die  Werthbestimmung  der  Kolanuss  und  des  Kolaex- 
traktes.  Vortrag  auf  der  Naturforscherversammlung  in  Braunschweig  gehalten. 
Pharmaceutische  Zeitung  42,  647-650  and  C  1897  II,  977. 

H.  Bnumer  and  H.  Leins. — Ueber  die  Trennung  und  quantitative  Bestim- 
mung des  Kaffeins  und  Theobromins.  Schweizer  Wochenschrift  fiir  Pharmacia 
36,  301-303  and  C  1898  II,  512. 

J.  Gadamer. — Ueber  Kaffeinbestimmungen  in  Tee,  Kaffee  und  Kola.  Archiv 
der  Pharmacie  237,  58-68  and  C  1899  I,  713. 

F.  Katz.— Ueber  die  quantitative  Bestimmung  des  Kaffeins.  Berichte  der 
Deutschen  pharmaceutischen  Gesellschaft  12,  250  (1902). 

I.  C.  C.  Keller's  Method. — Pour  120  grams  of  chloroform 
upon  6  griams  of  dry,  unbroken  tea  leaves^  in  a  wide-mouth 
separating  funnel.  In  a  few  minutes,  add  6  cc.  of  ammonium 
hydroxide  solution  (10  per  cent.  H3N),  and  at  frequent  intervals 
shake  vigorously  during  30  minutes.  Then  let  the  separating 
funnel  stand  at  rest,  until  the  solution  is  perfectly  clear  and  the 
tea  leaves  have  absorbed  all  the  water.  This  may  require 
3-6  hours,  or  even  longer,  depending  upon  the  variety  of  tea. 
Pass  100  grams  of  clear  chloroform  extract,  representing  5 
grams  of  tea,  through  a  small  filter  moistened  with  chloroform. 
Receive  the  filtrate  in  a  small,  weighed  flask  and  distil  the  chloro- 
form upon  the  water-bath  Pour  3-4  cc.  of  absolute  alcohol 
over  the  residue.  Heat  upon  the  water-bath  to  remove  alcohol 
and  expel  alcohol  vapor  with  a  hand  bellows.  In  a  few  minutes 
the  caffeine  will  be  dry  and  at  the  same  time  free  from  im- 

^  When  a  wide-mouth  separating  funnel  cannot  be  obtained,  triturate  the 
tea  leaves  somewhat,  solely  to  facilitate  their  removal  from  the  separating  funnel 
after  extraction.  Finely  powdered  tea  is  not  only  unnecessary',  but  even  ob- 
jectionable, because  the  extracts  have  a  much  deeper  color,  and  the  j^ield  of 
caffeine  is  not  increased. 


266  DETECTION    OF   POISONS 

prisoned  chloroform.  In  a  measure  also,  this  treatment  with 
alcohol  separates  caffeine  from  extraneous  chlorophyll.  The 
latter  adheres  to  the  bottom  and  side  of  the  flask,  whereas 
caffeine  forms  a  white  incrustation  upon  it.  Caffeine  thus 
obtained  is  usually  impure  from  small  quantities  of  ethereal 
oil,  fat,  vegetable-wax  and  principally  chlorophyll.  Conse- 
quently, it  must  be  purified.  Set  the  flask  upon  a  boiling  water- 
bath  and  pour  a  mixture  of  7  cc.  of  water  and  3  cc.  of  alcohol 
over  the  crude  caffeine,  which  upon  being  shaken  will  pass  into 
solution  almost  immediately.  Then  add  20  cc.  of  water, 
stopper  the  flask  and  shake  vigorously.  The  chlorophyll  will 
form  lumps  and  the  solution  will  filter  easily.  Pass  the  caffeine 
solution  through  a  small  filter  moistened  with  water,  wash  flask 
and  filter  with  10  cc.  of  water,  evaporate  the  total  filtrate  in 
a  weighed  glass  dish  to  dryness  upon  the  water-bath  and 
weigh  the  residue  of  nearly  pure  caffeine.  The  weight  of  this 
residue  multiplied  by  20  will  give  the  percentage  of  caffeine  in 
the  tea. 

Notes. — Ammonia  causes  tea  leaves  to  swell  considerably, 
and  at  the  same  time  combines  with  the  tannic  acid  present. 
Caffeine  is  set  free  and  dissolved  by  chloroform.  The  color  of 
the  chloroform  extract  depends  upon  the  variety  of  tea.  Black 
teas  (Pekoe,  Souchong  and  Congo)  give  clear,  pale  green  to 
yellowish  green  solutions.  Teas  not  so  black,  or  green  teas, 
give  darker  and  more  brownish  green  solutions.  In  assaying 
those  varieties  of  tea,  which  probably  contain  a  small  quantity 
of  caffeine,  take  12  grams  and  extract  with  150  grams  of 
chloroform.  C.  C.  Keller  has  shown  that  the  best  and  most 
expensive  varieties  of  tea  contain  most  caffeine.  The  average 
percentage  of  caffeine,  based  upon  50  assays  of  tea,  was  found 
to  be  3.06.  A  green  tea  gave  the  smallest  yield,  namely, 
1.78  per  cent,  of  caffeine;  and  a  Pekoe  tea  the  highest  yield, 
namely,  4.24  per  cent,  of  caffeine. 

J.  Gadamer  states  that  Keller's  method  of  estimating  caffeine 
in  tea  is  applicable  also  to  coffee  and  cola  preparations.  Keller's 
method  is  especially  useful  for  roasted  coffee.  The  caffeine, 
though  somewhat  brown,  is  always  sufficiently  pure. 


QUAN'I'I'I'ATIVK    lOS'lI  M  A'llON    OF    ALKALOIDS 


207 


2.  Hilger-Juckenack  Method. — Macerate  20  grams  of  finely 
ground  coffee,  or  triturated  tea,  with  900  grams  of  water  for 
several  hours  in  a  large  beaker  at  room  temperatures.  Then 
boil  thoroughly  and  replace  the  water  lost  by  evaporation. 
Raw  coffee  requires  3  hours  and  roasted  coffee  and  tea  1.5 
hours.  Cool  somewhat  (60  to  80°) 
and  add  75  grams  of  aluminum 
acetate  solution  (see  note,  page 
282)  and  gradually,  while  stirring, 
1.9  grams  of  acid  sodium  carbon- 
ate. Boil  about  5  minutes  and 
bring  the  total  weight  when  cold 
to  1020  grams.  Filter  750  grams 
(=15  grams  of  original  material) 
and  add  to  the  clear  filtrate  10 
grams  of  precipitated  and  pow- 
dered aluminium  hydroxide  and 
some  filter  paper  made  into  a 
magma  by  agitation  with  water. 
Stir  frequently  and  evaporate 
upon  the  water- bath.  Thoroughly 
dry  the  residue  in  an  air-closet  at 
100°,  and  extract  for  8-10  hours 
with  pure  tetrachloromethane 
(CCI4),  using  a  Soxhlet  apparatus 
(Fig.  23).  Tetrachloromethane, 
which  is  always  colorless,  is  finally 
distilled  and  the  residue  of  per- 
fectly white  caffeine  is  dried  at 
100°  and  weighed.  The  results 
thus  obtained  are  usually  accepted 
without  question.  But  if  an  absolutely  accurate  result  is 
required,  nitrogen  in  the  crude  caffeine  may  be  determined  by 
Kjeldahl's  method.  The  quantity  of  anhydrous  caffeine  is 
calculated  on  the  basis  of  this  analysis.  One  cc.  of  o.i  n-oxalic 
acid  represents  0.00485  gram  of  anhydrous  caft'eine.  Com- 
mercial tetrachloromethane  is  usually  impure  and  cannot  be 


Fig.   23. — Soxhlet  Apparatus. 


268  DETECTION   OF   POISONS 

used  directly  in  the  extraction.  It  should  be  shaken  3-4  times 
with  sodium  carbonate  solution,  then  several  times  with  water, 
dried  over  fused  calcium  chloride  and  distilled  fractionally. 
It  boils  at  76-77°. 

3.  Trillich-Goeckel  Modification  of  Hilger's  Method. — 
Exhaust  10  grams  of  finely  ground  coffee  with  water.  This 
will  require  3  extractions  with  boiling  water,  using  200  cc. 
portions  and  heating  each  for  30  minutes.  Combine  the  filtered 
extracts,  cool  and  dilute  to  495  cc.  Add  5  cc.  of  basic  lead  ace- 
tate solution,  shake  thoroughly,  filter  400  cc.  and  pass  hydrogen 
sulphide  through  the  filtrate.  Dilute  this  filtrate  to  500  cc, 
shake  thoroughly  and  again  filter  400  cc.  This  filtrate  will 
represent  6.4  grams  of  coffee.  Concentrate  this  filtrate 
(400  cc.)  upon  the  water-bath,  and,  after  addition  of  i  gram  of 
magnesium  oxide  and  sand,  evaporate  to  dryness.  Triturate 
the  residue  and  extract  for  30  hours  with  acetic  ether,  using  a 
Soxhlet  apparatus.  Evaporate  the  acetic  ether  extract  in  a 
Kjeldahl  flask,  or  distil  the  solvent,  and  determine  nitrogen  in 
the  residue  by  KjeldahFs  method.  One  gram  of  nitrogen  repre- 
sents 3.4643  grams  of  caffeine.  Crude  caffeine  is  easily  de- 
composed by  the  acid  used  in  the  Kjeldahl  process  and  by  mer- 
curic oxide.  Roasted  coffee  may  easily  give  too  much  caffeine 
by  this  method,  because  the  bases  formed  by  roasting  coffee, 
pyridine  for  example,  are  also  extracted  by  acetic  ether. 

4.  Trillich-Goeckel  Modification  of  Socolof's  Method. — 
Put  10  grams  of  finely  ground,  dried  coffee  into  a  separating 
funnel,  provided  with  a  plug  of  glass  wool  for  a  filter,  and 
moisten  with  ammonium  hydroxide  solution.  Let  the  mixture 
stand  for  30  minutes,  extract  for  12  hours  with  200  cc.  of  acetic 
ether  and  shake  frequently.  Filter,  and  wash  three  times  with 
50  cc.  portions  of  acetic  ether.  Distil  the  acetic  ether  upon  the 
water-bath  and  boil  the  residue  with  milk  of  magnesia.  Filter, 
and  evaporate  the  filtrate  to  dryness  upon  the  water-bath. 
Dissolve  the  residual  caffeine  in  acetic  ether  or  chloroform. 
Filter  this  solution  into  a  weighed  dish  or  Kjeldahl  flask. 
Evaporate  the  solvent  and  weigh  the  caffeine,  or  calculate  it 
from  the  percentage  of  nitrogen.     The  latter  method  is  the  more 


QUANTITATIVE    ESTIMATION   OF   AJ.KALOIDS  209 

accurate.  According  to  C.  Wolffs  the  residue  from  the  acetic 
ether  or  chloroform  extract  should  not  be  accepted  as  pure 
caffeine.  Determination  of  nitrogen  in  this  residue  by  Kjeldahl's 
method  is  the  most  reliable  way  of  estimating  caffeine  in  the 
extract. 

5.  E.  Katz's  Method  of  Estimating  Caffeine. — This  method 
is  based  upon  the  fact  that  chloroform  will  extract  caffeine 
quantitatively  from  a  solution  which  is  ammoniacal,  or  faintly 
acid  with  hydrochloric  acid. 

Shake  lo  grams  of  powdered  coffee,  or  tea,  for  30  minutes 
with  200  grams  of  chloroform  and  5  grams  of  ammonium 
hydroxide  solution.  When  the  liquid  has  settled,  filter  150 
grams  of  the  chloroform  solution  through  a  Sander's  filter 
which  will  give  a  perfectly  bright  filtrate  free  from  water.  Dis- 
til the  chloroform  completely  and  dissolve  the  residue  with 
gentle  heat  in  about  6  cc.  of  ether.  Add  20  cc.  of  0.5  per  cent, 
hydrochloric  acid  and,  in  an  assay  of  coffee,  also  0.2-0.5  gram 
of  solid  paraffine.  Evaporate  the  ether  and  filter  the  cold, 
aqueous  solution.  Wash  the  flask  and  filter  paper  a  few  times 
with  small  portions  of  0.5  per  cent,  hydrochloric  acid.  Finally 
extract  the  total  aqueous  hydrochloric  acid  solution  four  times 
with  20  cc.  portions  of  chloroform.  Distil  the  filtered  chloro- 
form extract,  dry  the  residue  and  weigh.  This  residue  will 
consist  of  nearly  pure  caffeine.  J.  Katz  found  the  following 
percentages  of  caffeine: 

Caffeine  Average 

Raw  Coffee  Beans 0.9  -1.27  per  cent.  1.14  per  cent. 

Dried  Cola  Nuts 1.51-1.94  per  cent.  1.68  per  cent. 

Black  Tea 2.51-3.56  per  cent.  3.07  per  cent. 

Guarana 2.83-4.74  per  cent.  4.0S  per  cent. 

J.  Katz  recommends  the  following  method  for  estimating 
caffeine  in  mate  or  Paraguay  tea: 

Treat  finely  triturated  tea  with  ammonium  hydroxide  solu- 
tion and  chloroform,  as  described  above,  and  dissolve  the  chloro- 
form residue  in  ether.  Add  water  to  the  ether  and  evaporate. 
Warm  the  aqueous  solution  10  minutes  upon  the  water-bath 

^  Zeitschrift  fiir  often tliche  Chemie  12,  186. 


270  DETECTION   OF   POISONS 

with  2  cc.  of  lead  hydroxide  suspended  in  water  (i  :2o).  If  it 
is  very  difficult  to  get  a  clear  filtrate  from  this  liquid,  add  a 
little  calcined  magnesium  oxide.  This  treatment  usually  gives 
a  filtrate,  which  is  perfectly  clear  when  cold,  and  but  shghtly 
colored.  Chloroform  extracts  quite  pure  caffeine  from  this 
solution.  By  this  method  mate  yields  0.3-1.6  per  cent,  of 
caffeine,  the  average  being  0.71  per  cent. 

6.  K.  Dieterich's  Method  of  Estimating  Total  Alkaloids 
(Caffeine  and  Theobromine)  in  Cola  Nuts. — Moisten  10  grams 
of  finely  grated  cola  nuts  with  a  little  water,  mix  with  10  grams 
of  granulated,  unslaked  lime  and  extract  with  chloroform  in  a 
Soxhlet  apparatus  for  45  minutes.  Evaporate  the  extract 
almost  to  dryness  and  dissolve  the  residue  with  gentle  heat 
in  20  cc.  of  normal  hydrochloric  acid.  Filter  and  dilute  to  100 
cc.  Add  ammonium  hydroxide  solution  in  large  excess  to  this 
filtrate,  shake  at  frequent  intervals  during  15  minutes  and 
extract  three  times  with  20  cc.  portions  of  chloroform.  Evapo- 
rate this  chloroform  solution  in  a  weighed  flask  and  dry  the 
residue,  which  usually  consists  of  perfectly  pure  caffeine,  at 
100°  to  constant  weight. 

This  method  may  also  be  used  in  estimating  caffeine  in 
Paraguay  tea.  Mix  the  finely  ground  material  with  unslaked 
Hme,  and  extract  with  chloroform,  in  a  Soxhlet  apparatus.  Tea 
gives  pure,  white  caffeine  free  from  chlorophyll. 

Estimation  of  Alkaloids  in  Ipecac 

Ipecac  has  been  shown^  to  contain  three  alkaloids : 

Cephaeline,  C28H40N2O4,         Emetine,  C30H44N2O4, 
Psychotrine. 

The  composition  of  the  last  alkaloid  is  unknown.  This  drug 
acts  as  an  expectorant  and  emetic,  because  of  cephaeline  and 
emetine.  Psychotrine  is  said  not  to  possess  these  properties. 
Therefore,  in  assaying  ipecac  for  medicinal  purposes,  only  the 
percentage  of  the  first  two  alkaloids  need  be  estimated.  The 
equivalent  weights  of  these  two  alkaloids  (cephaeline  234  and 

1  Frerichs  and  de  Fuentas  Tapis,  ArcMv  der  Pharmacie,  1902,  Heft  5  and  6. 


QUANTITATIVE     ESTIMATION    OF    ALKALOIDS  271 

emetine  248)  are  so  nearly  the  same,  that  the  mean  of  the  two 
(241)  may  be  used  as  the  factor. 

Procedure. — Put  6  grams  of  finely  powdered  root  in  a  dry 
Erlenmeyer  flask  and  shake  with  60  grams  of  ether.  'I'hen  add 
5  cc.  of  ammonium  hydroxide  solution,  or  5  cc.  of  sodium 
carbonate  solution  (1:3),  and  shake  frequently  during  an  hour. 
Add  10  cc.  of  water  and,  after  shaking  vigorously,  filter  50 
grams  of  the  ether  extract  into  a  small  flask.  Evaporate  half 
the  ether  upon  the  water-bath,  and  extract  the  remainder  in  a 
separating  funnel  with  10  cc.  of  o.i  n-hydrochloric  acid.  Pass 
the  acid  solution  through  a  small  filter  into  a  200  cc.  flask. 
Make  two  more  extractions  of  the  ether  with  10  cc.  portions  of 
water,  and  pass  these  through  the  same  filter.  Bring  the 
volume  of  the  acid  solution  to  100  cc,  and  then  add  enough 
ether  to  form  a  layer  about  i  cm.  thick  after  thorough  agitation. 
Add  5  drops  of  iodeosine  solution  (1:250),  and  titrate  excess  of 
hydrochloric  acid  with  o.i  n-potassium  hydroxide  solution. 
The  number  of  cc  of  0.1  n-hydrochloric  acid,  combined  with 
the  alkaloids,  multiplied  by  0.0241  gives  the  quantity  of  eme- 
tine and  cephseline  in  5  grams  of  ipecac. 

To  estimate  these  alkaloids  gravimetrically,  shake  vigorously 
the  ether  solution  of  the  alkaloids  (50  grams  =  5  grams  of  root) 
in  a  separating  funnel  with  5  cc.  of  dilute  hydrochloric  acid 
and  10  cc.  of  water.  Transfer  the  acid  solution  to  another 
separating  funnel.  Make  two  more  extractions  of  the  ether 
with  10  cc.  portions  of  water  and  add  these  to  the  acid  extract. 
Add  5  cc.  of  ammonium  hydroxide  solution  to  the  acid  extract 
and  shake  vigorously  with  50  grams  of  ether.  Remove  the  aque- 
ous layer  and  filter  40  grams  of  the  ether  solution  into  a  weighed 
flask.  Evaporate  the  ether  and  weigh  the  flask  after  drying 
for  an  hour  at  100°.  This  will  give  the  quantity  of  emetine  and 
cephfeline  in  4  grams  of  root. 

Test  for  Cephaeline. — This  reaction  is  very  characteristic  of 
this  alkaloid.  Froehde's  reagent  dissolves  pure  cephaeHne,  as 
the  free  base,  almost  without  color.  A  trace  of  hydrochloric 
acid,  or  better  sodium  chloride,  added  to  this  solution  produces 
an   intense   blue   color.     Pure   emetine   gives   no    color   with 


272  DETECTION   OF   POISONS 

Froehde's  reagent,  nor  when  sodium  chloride  is  added.     This 
test  for  cephaeline  may  be  made  with  the  ether  residue. 

The  method  of  estimating  alkaloids  in  ipecac,  prescribed  by  the  German 
Pharmacopoeia,  is  the  same  as  that  for  cinchona  bark.  Use  12  grams  of  finely 
powdered  root  dried  at  100°,  but  in  ascertaining  excess  of  acid  use  iodeosine, 
and  not  hasmatoxylin,  as  the  indicator.  Finally,  measure  with  a  pipette  50 
cc.  of  the  proper  solution  having  a  volume  of  100  cc,  place  in  a  200  cc.  flask 
and  add  about  50  cc.  of  water  and  enough  ether  to  make  a  layer  i  cm.  thick. 
Add  5  drops  of  iodeosine  solution  and  enough  o.oi  n-potassium  hydroxide 
solution,  shaking  thoroughly  after  each  addition,  to  give  the  lower  aqueous 
layer  a  pale  red  color.  This  should  require  not  more  than  20  cc.  of  alkaline 
solution. 

Estimation  of  Nicotine  in  Tobacco 

1.  R.  Kissling's^  Method. — First  remove  the  ribs  and  then 
cut  the  tobacco  leaves  into  small  pieces.  Dry  1-2  hours 
(50-60°),  and  then  reduce  to  a  uniform,  coarse  powder. 
Triturate  20  grams  of  this  powder  with  10  cc.  of  dilute,  alcoholic 
sodium  hydroxide  solution  (6  grams  of  sodium  hydroxide 
dissolved  in  40  cc.  of  water  and  60  cc.  of  95  per  cent,  alcohol). 
Transfer  this  moist  powder  to  a  paper  thimble  and  extract 
2-3  hours  with  ether  in  a  Soxhlet  apparatus.  Carefully  distil 
the  ether  solution  so  that  a  portion  of  the  solvent  remains.  Add 
50  cc.  of  very  dilute  sodium  hydroxide  solution  (4  grams  of 
sodium  hydroxide  in  1000  cc.  of  water)  to  the  residue  and  distil 
with  steam.  Begin  introducing  steam  after  the  nicotine  solu- 
tion has  been  boiling  several  minutes.  Collect  about  400  cc.  of 
distillate,  that  is  to  say,  continue  distillmg  until  the  distillate  is 
no  longer  alkaline.  Mix  well,  add  a  few  drops  of  rosolic  acid 
solution  to  the  distillate,  and  titrate  nicotine  with  o.i  n-sulphuric 
or  oxalic  acid  until  the  red  color  has  just  disappeared. 

Calculation. — Although  nicotine,  CioHuN2  (162),  as  adi-acid 
base  can  combine  with  two  equivalents  of  acid,  it  behaves  upon 
titration,  with  rosolic  acid  or  iodeosine  as  indicator,  as  if  it 
were  a  monacid  base  with  the  equivalent  weight  162.  1000  cc. 
of  0.1  n-acid  consequently  correspond  to  16.2  grams  of  nicotine. 

2.  C.  C.  Keller's^  Method. — Pour  60  grams  of  ether  and  60 

'^  Zeitschrif t  fiir  analytische  Chemie  34, 1731  and  21,  76. 

2  Berichte  der  Deutschen  pharmazeutischen  Gesellschaft  8,  145  (1898). 


QUANTITATIVE   ESTIMATION   OF   ALKALOIDS  273 

grams  of  petroleum  ether  over  6  grams  of  dry  tobacco  in  a  200 
cc.  Erlenmcyer  flask.  Add  10  cc.  of  20  per  cent,  aqueous 
potassium  hydroxide  solution  and  let  the  mixture  stand  half  an 
hour,  shaking  vigorously  at  frequent  intervals.  After  the  liquid 
has  stood  at  rest  3-4  hours,  pour  100  grams  of  ether  solution 
through  a  small,  plaited  filter  and  receive  the  filtrate  in  a  200  cc. 
Erlenmeyer  flask.  Nicotine  is  in  solution  together  with  a  Httle 
ammonia,  which  must  be  removed  before  titration.  By  means 
of  a  hand  bellows  and  a  glass  tube  reaching  to  the  bottom  of  the 
flask  force  a  current  of  air  through  the  solution,  so  that  there  is 
considerable  agitation.  It  requires  about  a  minute  and  a  half 
to  expel  all  ammonia.  At  the  same  time  8-10  grams  of  ether 
evaporate.  Add  10  cc.  of  alcohol,  a  drop  of  i  per  cent,  iodeo- 
sine  solution  and  10  cc.  of  water  to  the  ammonia-free  solution. 
Stopper  the  flask  and  shake  vigorously.  Nicotine  and  iodeo- 
sine  dissolve  in  the  water  which  has  a  red  color.  Add  a  slight 
excess  of  o.i  n-hydrochloric  acid,  enough  to  discharge  the  color, 
and  titrate  excess  of  acid  with  o.i  n-potassium  hydroxide  solu- 
tion. The  quantity  of  nicotine  in  tobacco  shows  a  wide  varia- 
tion and  ranges  from  0.6  to  4.8  per  cent. 

3.  J.  Toth's^  Method. — According  to  Toth  two  sources  of 
error  in  C.  C.  Keller's  method  lead  to  low  results.  An  aqueous 
potassium  hydroxide  solution  retains  variable  quantities  of 
nicotine  and  a  current  of  air  passed  through  an  ether  solution 
of  nicotine  volatilizes  some  of  this  alkaloid.  Therefore  Toth 
recommends  the  following  procedure: 

Mix  6  grams  of  air-dried  tobacco  with  10  cc.  of  20  per  cent, 
sodium  hydroxide  solution  in  a  porcelain  dish.  Add  gypsum 
until  the  mixture  is  like  powder.  Extract  thoroughly  with 
100  cc.  of  ether-petroleum  ether  mixture  (1:1)  and  after  i  hour 
pipette  off  as  quickly  as  possible  25  cc.  of  the  solvent.  Add 
40-50  cc.  of  water,  i  drop  of  iodeosine  solution  and  an  excess  of 
O.I  n-sulphuric  acid.  Determine  excess  of  acid  by  titration 
with  O.I  n-sodium  hydroxide  solution.  The  ether-petroleum 
ether  mixture  takes  up  at  most  0.0005  gram  of  ammonia. 

^  Chemisches  Zentralblatt,  1901,  i,  973. 
18 


274  DETECTION   OF  POISONS 

Estimation  of  Hydra  stine  in  Flixid  Extract  of  Hydrastis 

(German  Pharmacopoeia) 

Evaporate  15  grams  of  fluid  extract  of  hydrastis  to  about  5  grams  in  a  weighed 
dish  upon  the  water-bath,  and  wash  the  residue  into  an  Erlenmeyer  flask  with 
about  10  cc.  of  water.  Add  10  grams  of  petroleum  ether,  50  grams  of  ether  and  5 
grams  of  ammonium  hydroxide  solution.  Let  the  mixture  stand  an  hour,  shaking 
vigorously  at  frequent  intervals.  Then  pass  50  grams  of  the  clear  ether  solution 
through  a  dry  filter  into  a  separating  funnel.  Add  10  cc.  of  a  mixture,  composed 
of  I  part  of  hydrochloric  acid  and  4  parts  of  water,  and  shake  the  solution 
vigorously  several  minutes.  When  the  liquids  have  separated  clear,  run  the 
acid  solution  into  an  Erlenmeyer  flask.  Make  two  more  extractions  of  the  ether 
with  5  cc.  portions  of  water  containing  a  few  drops  of  hydrochloric  acid,  and  add 
these  to  the  first  extract.  Add  to  the  total  extract  excess  of  ammonium  hydroxide 
solution  and  50  grams  of  ether.  Let  the  mixture  stand  an  hour,  shaking  vigor- 
ously at  frequent  intervals.  Pass  40  grams  of  the  clear  ether  solution  through 
a  dry  filter  and  collect  the  filtrate  in  a  weighed,  dry  flask.  Distil  the  ether, 
dry  the  residue  at  100°  and  weigh  when  cold.  The  residue  should  weigh  at 
least  0.2  gram. 

Notes. — Additional  information  about  hydrastine  is  given 
on  page  112.  Ammonia,  added  to  an  aqueous  solution  of  the 
residue  from  hydrastis  extract  (15  grams),  sets  the  alkaloids, 
hydrastine  and  berberine,  free  from  their  salts.  The  ether- 
petroleum  benzine  mixture  dissolves  hydrastine  but  not  ber- 
berine, the  latter  being  nearly  insoluble  in  this  mixed  solvent. 
But  phytosterin,  which  is  always  present  in  hydrastis  extract,  is 
dissolved.  Only  50  grams  (=  hydrastine  in  12.5  grams  of 
extract)  of  the  original  60  grams  of  ether-petroleum  benzine 
mixture  are  used.  Hydrastine  is  extracted  from  the  solvent  by 
agitation  with  dilute  hydrochloric  acid  and  dissolved  in  the 
acid  solution  as  hydrochloride.  The  alkaloid  is  then  precipi- 
tated from  the  acid  solution  by  ammonia  and  dissolved  in  50 
grams  of  ether: 

C21H21NO6.HCI  +  (H4N)0H  =  C21H21NO6  +  H2O  +  (H4N)C1. 

Hydrastine  Hydrastine 

hydrochloride 

The  hydrastine  in  40  grams  of  the  ether  solution  (=10  grams 
of  original  extract)  is  finally  weighed.  Good  extract  of  hydras- 
tis should  contain  2-2.5  P^^  cent,  of  hydrastine. 

When  the  ether-petroleum  benzine  solution  of  hydrastine  and 
phytosterin  is  extracted  with  dilute  hydrochloric  acid,  the 
alkaloid  passes  into  the  acid  solution  free  from  phytosterin. 


QUANTITATIVE   ESTIMATION    OF    ALKALOIDS  275 

Estimation  of  Berberine. — This  alkaloid  has  only  a  slight  physiological  action. 
To  dclcrminc  ap))r()ximalcly  the  quantity  present  in  hydrastis  extract,  add  20 
grams  of  dilute  sulphuric  acid  (i :  5)  to  10  grams  of  the  extract  and  let  the  mixture- 
stand  for  24  hours  at  as  low  a  temperature  as  possible.  Crystallization  of  ber- 
berine as  the  difficultly  soluble  acid  sulphate,  C20H17NO4.H2SO4,  is  almost  com- 
plete. Filter  in  a  Gooch  crucible  with  suction,  washing  first  with  a  little  water 
containing  sulphuric  acid  and  then  with  pure  water.  Dry  at  100°  to  constant 
weight.     (E.  Schmidt.) 

W.  Meine^  has  found  that  the  crystalline  deposit,  frequently  seen  in  hydrastis 
extract,  consists  mostly  of  berberine  mixed  with  a  little  phytosterin.  This 
deposit  is  said  to  contain  only  traces  of  hydrastinc. 

Picrolonate  Method  of  Estimating  Hydrastine  in  Hydrastis  Root  and 

Extract 

(H.  Matthes  and  O.  Rammstedt)^ 

The  German  Pharmacopoeia  requires  the  estimation  of 
hydrastine  but  not  of  the  physiologically  inert  substances^ 
berberine  and  phytosterin,  also  present  in  hydrastis  prepara- 
tions. Ether-petroleum  benzine  mixture,  used  as  a  solvent, 
dissolves  phytosterin  and  hydrastine  but  not  berberine.  Di- 
lute hydrochloric  acid  extracts  hydrastine  but  leaves  phytos- 
terin in  the  ether  mixture. 

Estimation  of  hydrastine  by  means  of  picrolonic  acid  appears 
simpler  than  by  the  method  of  the  Pharmacopoeia,  because 
picrolonic  acid  does  not  precipitate  phytosterin  and  therefore 
hydrastine  is  not  mixed  with  this  impurity.  Matthes  and 
Rammstedt  obtained  nearly  pure  hydrastine  picrolonate  from 
hydrastis  extract,  melting  at  220-225°.  The  picrolonate  pre- 
pared from  pure  hydrastine,  C21H21NO6.C10H8N4O5,  melts  at 
225°. 

I.  In  Fluid  Extract  of  Hydrastis.^Evaporate  15  grams  of 
fluid  extract  to  about  5  grams  in  an  Erlenmeyer  flask  upon  the 
water-bath.  Add  10  cc.  of  water;  10  grams  of  petroleum 
benzine,  50  grams  of  ether  and  5  grams  of  ammonium  hydroxide 
(10  per  cent.  NH3).  Shake  vigorously  for  10  minutes.  After 
the  mixture  has  stood  for  20  minutes,  pour  40  grams  of  the  ether- 

1  Zeitschrift  des  allgemeinen  osterreichischen  Apotheker-Vereins  55,  494. 
-  Further  information  about  picrolonic  acid  and  its  use  in  precipitating  alkaloids 
is  given  on  page  246. 


276  DETECTION    OF   POISONS 

benzine  extract  through  a  double,  creased  filter  and  evaporate 
about  one-half  in  a  beaker.  Then  add  lo  cc.  of  o.i  n-picrolonic 
acid  solution.  After  24  hours  collect  the  hydrastine  picrolonate 
in  a  weighed  Gooch  crucible,  wash  with  2  cc.  of  an  alcohol-ether 
mixture  (1:3),  dry  for  30  minutes  at  105°  and  weigh. 

2.  In  Hydrastis  Root. — Shake  6  grams  of  powdered  root 
vigorously  for  30  minutes  with  50  grams  of  ether,  10  grams  of 
petroleum  ether  and  6  grams  of  ammonium  hydroxide  (10  per 
cent.  NH3).  Then  add  6  grams  of  water  and  shake  until  the 
upper  layer  of  liquid  is  clear.  Quickly  filter  50  grams  of  the 
ether-petroleum  benzine  extract  and  evaporate  about  one-half 
in  a  beaker.  Then  add  5  cc.  of  o.i  n-picrolonic  acid.  After  24 
hours  filter  the  picrolonate  precipitate  and  wash  with  i  cc.  of 
alcohol-ether  mixture  (1:3).  Otherwise  the  procedure  is  the 
same  as  described  in  i . 

In  the  calculation  use  the  formula  of  hydrastine  picrolonate 
(Mol.  Wt.  647)  given  above. 

Estimation  of  Morphine  in  Opium  and  Pharmaceutical  Preparations 

(German  Pharmacopoeia) 

In  Opium. — Triturate  6  grams  of  rather  finely  powdered  opium  with  6  grams  of 
water.  Wash  the  mixture  into  a  weighed,  dry  flask  with  water  and  add  enough 
more  of  this  solvent  to  bring  the  weight  to  54  grams.  Shake  frequently  and  let 
the  mixture  stand  an  hour.  Pour  upon  a  piece  of  dry  linen  and  express  the 
liquid.  Pass  42  grams  of  this  extract  through  a  dry,  plaited  filter  (10  cm.  in  di- 
ameter) into  a  dry  flask.  Add  2  grams  of  sodium  salicylate  solution  (i :  2)  to  this 
filtrate  and  shake  vigorously.  Filter  36  grams  of  the  clear  solution  through  a  dry, 
plaited  filter  (10  cm.  in  diameter)  into  a  small  flask.  Mix  this  filtrate  by  gentle 
agitation  with  10  grams  of  ether,  and  add  also  5  grams  of  a  mixture  consisting  of 
17  grams  of  ammonium  hydroxide  solution  and  83  grams  of  water.*  Stopper  the 
flask,  shake  vigorously  for  10  minutes  and  let  the  mixture  stand  at  rest  24  hours. 
Then  decant  the  ether  layer  as  completely  as  possible  upon  a  smooth  filter 
(8  cm.  in  diameter).  Add  10  grams  more  of  ether  to  the  residual,  aqueous  liquid 
in  the  flask,  shake  gently  for  a  few  minutes  and  again  pour  the  ether  layer  upon 
the  filter.  Then  after  all  the  ether  solution  has  passed  through,  pour  the 
aqueous  solution  upon  the  filter,  and  disregard  crystals  adhering  to  the  side  of 
the  flask.  Wash  filter  and  flask  three  times  with  5  cc.  portions  of  water  satu- 
rated with  ether.  When  the  filter  has  drained  thoroughly,  dry  the  morphine 
crystals  and  dissolve  in  25  cc.  of  0.1  n-hydrochloric  acid.  Pour  this  solution 
into  a  100  cc.  flask,  carefully  wash  filter  and  flask  with  water  and  finally  dilute 
the  solution  to  100  cc.     Measure  50  cc.  of  this  solution  into  a  200  cc.  flask,  add 


QUANTITATIVE   ESTIMATION    OF   ALKALOIDS  277 

50  cc.  of  water  and  enough  ether  to  form  a  layer  i  cm.  thick.  Add  5  drops  of 
iodeosine  solution  and  enough  o.i  n-potassium  hydroxide  solution,  shaking  vigo- 
rously after  each  addition,  to  produce  a  pale  red  color  in  the  lower  aqueous  layer.' 

Notes  and  Calculation. — Most  of  the  opium  alkaloids  are 
combined  with  meconic  (see  page  205)  and  suli)huric  acids. 
Ammonium  hydroxide,  added  to  an  aqueous  opium  extract, 
sets  the  alkaloids  free  from  their  salts: 

(CitHiaNOz  oh 

C6H02(OH)(COOH)2  +  2(H4N)OH  =  2C17H19NO3  +  2H2O  +  CsHOz 

II 
(COONHOz 

Morphine  meconate  Morphine  Ammonium 

meconate 

The  ether  used  dissolves  all  opium  alkaloids  except  morphine 
which  having  once  become  crystalline  is  insoluble  in  this 
solvent.  Saturated  sodium  salicylate  solution  precipitates 
resinous  and  greasy  substances  from  the  filtered  aqueous  opium 
extract  and  also  narcotine  which  next  to  morphine  is  present  in 
opium  in  largest  quantity. 

The  moprhine  from  6  grams  of  opium  is  in  54  grams  of  filtered 
aqueous  extract.  After  the  second  filtration  only  36  grams  of 
this  extract  are  used  (=  morphine  from  4  grams  of  opium). 
Morphine,  precipitated  by  ammonium  hydroxide  from  these  36 
grams  of  extract,  is  dissolved  in  25  cc,  of  o.i  n-hydrochloric  acid 
as  hydrochloride,  CnHigNOs.HCl.  This  solution  is  then  diluted 
to  100  cc,  and  excess  of  acid  in  50  cc,  of  this  hydrochloric  acid 
solution  (=  morphine  from  2  grams  of  opium)  is  determined. 

Morpine  being  a  monacid  base  has  the  same  molecular  and 
equivalent  weights  =  C17H19NO3  =  285.  Therefore  1000  cc. 
of  O.I  n-hydrochloric  acid  =  28.5  grams  of  morphine. 

Example. — Titration  with  0.1  n-potassium  hydroxide  solution  has  shown  that 
there  are  4.1  cc.  of  0.1  n-hydrochloric  acid  in  50  cc.  of  the  hydrochloric  acid 
solution  of  morphine.  There  remain  therefore  12.5  —  4.1  =  8.4  cc.  of  the  0.1 
n-acid  originally  present  now  combined  mth  the  morphine  from  2  grams  of  opium. 
According  to  the  proportion 

Cc.  O.I  n-HCl  :  Grams   morphine 

1000  :  28  .5  =  8.4  :  X     (x  =  0.2394) 

^  The  German  Pharmacopoeia  demands  that  not  more  than  5.4  cc.  nor  less  than 
4.1  cc.  of  O.I  n-alkaline  hydroxide  solution,  corresponding  to  a  morphine-content 
of  IC-12  per  cent,  shall  be  used  to  produce  this  color. 


278  DETECTION   OF   POISONS 

8.4  cc.  of  O.I  n-acid  correspond  to  0.2394  gram  of  morphine.  Consequently  the 
opium  contains  50  X  0.2394  =  11.97  per  cent,  of  morphine.  This  is  the  maxi- 
mum quantity  of  morphine  allowed  in  opium  by  the  German  Pharmacopoeia. 

2.  In  Extract  of  Opium.  "Dissolve  3  grams  of  opium  extract  in  40  grams  of 
water,  add  2  grams  of  sodium  salicylate  solution  (i:  2),  shake  vigorously,  pass  30 
grams  of  clear  solution  through  a  dry  filter  (10  cm.  in  diameter)  and  collect 
in  a  dry  flask.  Mix  this  filtrate  with  10  grams  of  ether  by  rotating  the  flask  and 
add  also  5  grams  of  a  mixture  of  1 7  grams  of  ammonium  hydroxide  and  83  grams 
of  water."  Continue  the  assay  as  directed  above  in  i  (Opium)  from  the  point 
marked  with  an  asterisk. 

Calculation. — Only  2  of  the  3  grams  of  opium  extract  weighed  are  used  in  the 
determination,  since  only  30  grams  of  the  original  45  grams  of  solution  (3  grams 
of  extract  +  2  grams  of  sodium  salicylate  solution  +  40  grams  of  water)  are 
filtered.  The  morphine  obtained  from  these  2  grams  of  extract  is  dissolved  in  25 
cc.  of  o.  I  n-hydrochloric  acid  and  the  volume  is  then  brought  to  100  cc.  The  titra- 
tion uses  50  cc.  of  this  solution  which  contains  the  morphine  from  i  gram  of 
opium  extract. 

Example. — If  5.5  cc.  of  o.i  n-potassium  hydroxide  solution  were  required  to 
neutralize  the  excess  of  o.i  n-hydrochloric  acid  in  the  50  cc.  of  solution,  then 
12.5  —  5.5  =  7  cc.  of  O.I  n-hydrochloric  acid  are  combined  with  morphine.  Ac- 
cording to  the  proportion 

Cc.  O.I  n-HCl  :  Grams    morphine 

1000  :  28.5  =  7  :  X     (x  =  0.1995) 

7  cc.  of  O.I  n-acid  correspond  to  0.1995  gram  of  morphine.  Therefore  this 
■quantity  of  alkaloid  is  in  i  gram  of  extract.  Consequently  the  opium  extract 
■contains  19.95  P^r  cent,  of  morphine. 

The  German  Pharmacopoeia  requires  that  not  more  than  6.5  cc.  nor  less  than 
.5.5  cc.  of  O.I  n-potassium  hydroxide  solution  shall  be  used  to  produce  a  pale 
red  color  in  the  aqueous  layer,  corresponding  to  a  morphine  content  of  17. 11  to 
19.95  per  cent. 

3.  In  Wine  of  Opium  and  Tinctiu-e  of  Opium.^ — "Evaporate  about  50  grams 
of  either  preparation  in  a  weighed  dish  to  15  grams,  add  water  until  the  weight 
is  38  grams  and  also  2  grams  of  sodium  salicylate  solution  (i  :  2).  Shake  vig- 
orously and  pass  32  grams  of  clear  solution  through  a  dry  creased  filter  (10  cm. 
in  diameter)  into  a  dry  flask.  Mix  this  filtrate  with  10  grams  of  ether  by  rotating 
the  flask  and  add  also  5  grams  of  a  mixture  of  1 7  grams  of  ammonium  hydroxide 
solution  and  83  grams  of  water."  Continue  the  assay  as  directed  above  in  i 
(Oj)ium)  from  the  point  marked  with  an  asterisk. 

Calculation. — Only  32  grams  of  the  40  grams  of  clear  liquid  (38  grams  of  evapo- 
rated opium  tincture  +  2  grams  of  sodium  salicylate  solution)  are  used  for  the 
morphine  determination.  These  correspond  to  40  grams  of  the  original  opium 
preparation.  The  morphine  from  this  quantity  of  solution  is  dissolved  in  25 
cc.  of  O.I  n-hydrochloric  acid  and  the  volume  brought  to  100  cc.  Excess  of  acid 
in  50  cc.  of  this  solution  is  determined  by  titration.  These  50  cc.  contain  the 
morphine  from  20  grams  of  the  opium  preparation. 

Example. — If  4.2  cc.  of  0.1  n-potassium  hydroxide  solution  are  required  for 


QUAN'I'I'I'ATIVE    ESTIMATION    OF    ALKALOIDS  279 

50  cc.  of  mori)Iiinc  liydrocliloridc  soliilion,  llion  12.5  —4.2  =  8.3  cc.  of  o.i  n- 
hydrochloric  acid  have  combined  willi  mor|)liinc.     According  to  the  proportion 

Cc.  0.1  n-IICl  :  Grams   morphine 

1000  :  28.5  =  8.3:x     (x  =  0.23655) 

20  grams  of  the  opium  preparation  contain  0.23655  gram  of  morphine,  corre- 
sponding to  a  morplaine  content  of  1.18  per  cent. 

The  German  Pharmacopoeia  requires  that  not  more  than  5.5  cc.  nor  less  than 
4.2  cc.  of  0.1  n-potassium  hydroxide  solution  shall  be  used  to  produce  a  pale 
red  color  in  the  aqueous  liquid,  corresponding  to  a  morphine  content  in  Wine 
of  Opium  and  Tincture  of  Opium  of  i.o  to  1.18  per  cent. 


Estimation  of  Pilocarpine  in  Jaborandiim  Leaves^ 

1.  G.  Fromme's-  Method. — Extract  15  grams  of  rather 
finely  powdered  jaborandum  leaves  with  150  grams  of  chloro- 
form and  15  grams  of  ammonium  hydroxide  solution  (10  per 
cent.  NH3),  shaking  frequently  for  30  minutes.  Filter  this 
mixture  through  a  large,  smooth  paper,  covering  the  funnel 
with  a  glass  plate.  As  soon  as  the  chloroform  drops  slowly,  add 
a  little  water  and  filtration  will  become  more  rapid.  After 
collecting  a  full  100  grams  of  filtrate,  add  about  i  gram  of  water, 
shake  vigorously  and  set  aside.  The  water  takes  up  fine 
particles  of  powder  that  may  have  passed  through  the  paper, 
leaving  the  chloroform  solution  quite  clean.  After  i  hour  weigh 
100  grams  of  chloroform  solution  (=  alkaloids  in  10  grams  of 
jaborandum  leaves). 

Fromme  directs  extracting  these  100  grams  of  chloroform 
solution  successively  with  30,  20  and  10  cc.  of  i  per  cent,  hydro- 
chloric acid  which  dissolves  pilocarpine  (and  isopilocarpine)  as 
hydrochlorides.  Extract  this  acid  solution  first  with  20  cc.  of 
ether,  to  remove  fat  and  resin.  Then  add  an  excess  of  ammonia 
and  extract  the  free  alkaloids  successively  with  30,  20  and  10  cc. 
of  chloroform.  Pour  the  combined  chloroform  extracts  through 
a  dry  filter,  evaporate  in  a  weighed  flask,  dry  the  residue  at  100° 
and  weigh. 

2.  Matthes  and    Rammstedt's^   Method. — Evaporate    100 

'  Further  information  about  pilocarpine  is  given  on  page  210. 
'^  Caesar  and  Loretz,  Geschaftsbericht  1901,  27. 
2  See  page  246. 


280  DETECTION   OF   POISONS 

grams  of  chloroform  solution  obtained  above  in  a  beaker  to 
about  20  cc;  Add  first  3  cc.  of  o.i  n-picrolonic  acid  and  then 
60  cc.  of  ether.  After  24  hours  collect  the  precipitate  of  pilo- 
carpine picrolonate  in  a  weighed  Gooch  crucible,  wash  with  i  cc. 
of  alcohol-ether  mixture  (1:3),  dry  at  110°  and  weigh.  Pilo- 
carpine picrolonate  thus  obtained  (=  pilocarpine  from  10  grams 
of  jaborandum  leaves),  CHH16N2O2.C10H8N4O5  (Mol.  Wt.  472) 
melts  at  200-205°. 

Piperine  in  Pepper 

Black  pepper  is  the  dried,  vmripe  fruit  of  the  pepper  plant,  Piper  nigrum  L., 
whereas  the  ripe  fruit  deprived  of  its  outer  covering  is  the  white  pepper  of  com- 
merce. The  actual  constituents  of  pepper  are  piperine,  an  ethereal  oil  (oil  of 
pepper)  and  a  resin  called  chavicine.     In  rather  large  doses  pepper  is  toxic.^ 

Preparation  of  Piperine. — Extract  finely  divided  white  pepper  with  90  per  cent, 
alcohol  and  distil  the  latter  from  the  extracts.  Treat  the  residue  with  cold 
potassium  hydroxide  solution  which  dissolves  the  resin  but  not  the  piperine. 
Wash  the  residual  piperine  with  water  and  crystallize  from  hot  alcohol,  using 
animal  charcoal  to  remove  color.     White  pepper  contains  7-8  per  cent,  of  piperine. 

Piperine,  C17H19NO3,  crystallizes  in  colorless,  shining,  rectangular,  monoclinic 
prisms  melting  at  1 28-1 29°.  When  pure  it  is  almost  tasteless  but  impure  piperine 
has  a  sharp,  burning  taste.  It  is  nearly  insoluble  in  water,  freely  soluble  in  alco- 
hol and  also  soluble  in  ether,  benzene  and  chloroform.  Piperine  is  a  very  weak 
base,  dissolving  in  dilute  mineral  acids  with  almost  as  much  difficulty  as  in  pure 
water.  A  solution  of  piperine  in  concentrated  sulphuric  acid  has  a  ruby  color, 
soon  changing  to  dark  brown  and  gradually  to  greenish  brown  and  fading  upon 
addition  of  water.  Concentrated  nitric  acid  converts  piperine  into  an  orange 
red  resin  soluble  with  blood-red  color  in  dilute  potassium  hydroxide  solution. 
The  constitution  of  piperine  is  known. 

Prolonged  heating  with  alcohoHc  potassium  hydroxide  solution  decomposes 
piperine  into  the  potassium  salt  of  piperic  acid  and  piperidine : 

CH  =  CH.CO.NCeHio  CH  =  CH.COOK 

1  +  KOH  =  CbHioNH  -f-  I 

CH  =  CH.CeHsCCOaHa)  Piperidine  CH  =  CH.CeHsCCOaHz) 

Piperine  Potassium  piperate 

Rugheimer^  synthesized  piperine  by  putting  together  these  two  products, 
Piperic  acid  was  first  converted  into  its  chloride  by  means  of  phosphorus  penta- 

^  R.  Kobert  ("  Intoxikationen ")  mentions  a  case  where  a  teaspoonful  of 
pepper  was  given  to  each  of  three  young  pigs.  There  was  severe  inflammation 
of  the  gastro-intestinal  tract  in  all  three  animals  and  two  died.  The  toxic  action 
of  pepper  is  attributed  to  piperine,  since  the  ethereal  oil  according  to  Kobert  does 
not  take  part  in  the  toxic  action  due  to  absorption. 

2  Berichte  der  Deutschen  chemischen  Gesellschaft  15,  1390  (1882). 


QUANTITATIVE   ESTIMATION    OF   ALKALOIDS  281 

chloride.     Piperyl  chloride  was  then  condensed  in  benzene  solution  with  piperi- 
dine: 

CH  =  CH.COOPI  CH  =  CH.C0C1 

I  +  PCl6  =   i  +  POCI3  +  HCl 

CH  =  CH.CoHsCCOJIa)  CH  =  Cri.CoIIsCCOalTz) 

CH  =  CH.C0C1  CH  =  CH.CO.NC6ir,o 

I  +  HNCbPIio  =    I  +  HCl. 

CH  =  CH.CflH3(C02H2)  .  .  CH^CH.CeHaCCOsHz) 

Piperyl  chloride  Piperidine  Piperine 

On  the  basis  of  the  known  structure  of  piperic  acid  and  piperidine,  piperine 
must  have  the  following  constitution: 

H  H2 

C  C 

/\  /\ 

O— C      CH  HoC      CHo 


H2C 


0— C      CH  H2C      CH2 

\/  \/ 

C  N 

I  I 

'— CH=CH.CH=CH.CO— ' 


Estimation  of  Piperine  in  Pepper 


1.  J.  Koenig's  Method.- — Exhaust  10-20  grams  of  pepper, 
ground  as  finely  as  possible,  in  a  Soxhlet  apparatus  with  strong 
ethyl  or  methyl  alcohol,  or  petroleum  ether.  Distil  the  alcohol 
or  petroleum  ether.  The  residue  consists  of  piperine  and  resin. 
Shake  this  residue  with  cold  potassium  or  sodium  carbonate 
solution  to  dissolve  the  resin.  Filter  from  undissolved  piperine 
and  wash  the  latter  with  cold  water.  Dissolve  in  alcohol  or 
petroleum  ether,  evaporate  the  filtered  solution  in  a  weighed 
flask  or  dish  and  dry  the  residue  at  100°  to  constant  weight. 

To  determine  the  resin  in  pepper  at  the  same  time,  filter  the 
potassium  or  sodium  carbonate  solution  from  crude  piperine  and 
■  add  hydrochloric  acid  to  the  filtrate.  Filter  the  precipitated 
resin,  redissolve  in  alcohol,  evaporate  the  solvent  and  dry  the 
residue  to  constant  weight. 

2.  Cazeneuve  and  Caillot's  Method. — Add  enough  water  to 
make  a  thin  mixture  of  powdered  pepper  with  twace  its  weight 
of  slaked  lime  and  stir  well.  Boil  in  a  porcelain  dish,  dr}-  thor- 
oughly upon  the  water-bath  and  then  extract  with  ether  in  a 
Soxhlet  apparatus.     Distil  the  ether  in  a  weighed  flask  and 


282  DETECTION   OF   POISONS 

dry  the  residue  of  piperine  at  ioo°  to  constant  weight.  To 
obtain  purecrystalHne  piperine,  dissolve  the  residue  from  the 
ether  distillation  in  the  least  possible  volume  of  boiling  alcohol, 
surround  the  solution  with  ice,  collect  the  piperine  upon  a 
weighed  filter  and  dry  at  ioo°  to  constant  weight.  This 
purification  of  piperine  is  attended  with  more  or  less  loss  and 
consequently  the  result  is  only  approximately  correct. 

Estimation  of  Santonin  in  Wormseedi 

I.  K.  Thaeter's^  Method. — Extract  lo  grams  of  crushed 
wormseed  in  a  Soxhlet  apparatus  with  ether  for  12  hours. 
Distil  the  ether  and  boil  the  residue  for  an  hour  with  5  grams 
of  lime  and  about  300  cc.  of  water.  Replace  water  lost  by 
evaporation.  Filter  while  hot  and  wash  the  residue  with 
water.  Faintly  acidify  the  filtrate  with  sulphuric  acid  and 
warm  gently  until  santonin  crystals  begin  to  form.  Then  add 
100  grams  of  aluminium  acetate  solution,^  heat  the  mixture 
to  boiling  and  finally  evaporate  to  dryness  upon  the  water- 
bath.  Mix  the  finely  powdered  residue  with  3  grams  of  mag- 
nesium oxide,  moisten  this  mixture  with  a  little  water  and  again 
bring  quickly  to  dryness.  Powder  the  residue  as  finely  as  pos- 
sible, dry  at  105°  and  extract  in  a  Soxhlet  apparatus  with  an- 
hydrous, acid-free  ether  for  5  hours.  Santonin  is  deposited 
upon  distilling  the  ether  as  a  faintly  yellowish  residue  which 
is  then  dried  at  100°  to  constant  weight. 

Remarks. — When  wormseed  is  heated  with  lime,  santonin  passes  into  solution 
as  calcium  santonate,  and  at  the  same  time  resinous  substances  are  saponified. 

^  Wormseed  (Flores  cinas)  consists  of  the  unexpanded  flower -heads  of  Artemisia 
cina  which  are  3-4  mm.  in  length. 

^  Archiv  der  Pharmacie  237,  626-632  (1899)  and  238,  383-387  (1900). 

'  Dissolve  300  parts  of  aluminium  sulphate  in  800  parts  of  water;  add  acetic 
acid  (sp.  gr.  1.041)  360  parts;  triturate  calcium  carbonate  130  parts  with  200 
parts  of  water,  and  add  this  mixture  slowly  and  with  continued  stirring  to  the 
first  solution;  set  the  whole  aside  for  24  hours  without  applying  heat,  and  stir 
occasionally;  then  strain,  press  the  precipitate  without  washing  it  and  filter  the 
liquid.  It  is  a  clear,  colorless  liquid,  having  the  sp.  gr.  1.044  to  1.046,  a  faint 
odor  of  acetic  acid,  an  acid  reaction,  and  a  sweetish,  astringent  taste.  National 
Dispensatory. 


QUANTITA'I'IVE    ESTIMATION    OF    ALKALOIDS  283 

Dilute  sulphuric  acid  liberates  first  santonic  acid  which  passes  at  once  into  its 
inner  anhydride,  santonin.  Basic  alurninium  acetate,  produced  by  boiling, 
precipitates  resinous  and  colored  substances.  Finally,  magnesium  oxide  serves 
to  neutralize  free  acetic  acid.  Under  the  conditions,  practically  no  magnesium 
santonate  is  formed.  Thaeter  obtained  88  to  92  per  cent,  of  the  santonin  present. 
Wormseed  contains  about  2.5  per  cent,  of  santonin. 

2.  J.  Katz's"^  Method. — Extract  lo  grams  of  coarsely  pow- 
dered wormseed  in  a  Soxhlet  apparatus  with  ether  for  2  hours. 
Distil  the  ether.  There  usually  remains  a  dark  green  resin 
weighing  1.5-2  grams.  Boil  this  residue  15-30  minutes,  under  a 
reflux  condenser,  with  5  grams  of  crystallized  barium  hydroxide 
dissolved  in  100  cc.  of  water.  Cool  and,  without  filtering, 
render  the  solution  acid  to  litmus  with  carbon  dioxide.  Filter 
immediately  with  a  pump  to  remove  barium  carbonate,  and 
wash  the  precipitate  twice  with  20  cc.  portions  of  water.  Evapo- 
rate the  pale  yellow  solution  to  about  20  cc.  in  a  dish  upon  the 
water-bath.  Add  10  cc.  of  12.5  per  cent,  hydrochloric  acid, 
continue  heating  upon  the  water-bath  exactly  2  minutes  longer 
and  pour  the  solution  into  a  separating  funnel.  Dissolve 
santonin  crystals  left  in  the  dish  in  about  20  cc.  of  chloroform. 
Pour  this  solution  into  the  separating  funnel  and  shake  thor- 
oughly. When  the  solutions  have  separated  clear,  withdraw 
the  chloroform  solution  and  pass  it  through  a  dry  filter.  Wash 
dish,  separating  funnel  and  filter  2-3  times  with  10  cc.  portions 
of  chloroform.  Distil  the  chloroform  and  boil  the  residue  10 
minutes,  under  a  reflux  condenser,  with  50  cc.  of  15  per  cent, 
alcohol.  Filter  while  hot  into  a  weighed  flask,  and  wash  flask 
and  filter  twice  with  15  cc.  portions  of  15  per  cent,  boifing 
alcohol.  Cover  the  flask  and  set  aside  in  the  cold  24  hours. 
Weigh  flask  and  contents  and  pass  the  latter  through  a  weighed 
filter,  disregarding  the  milky  appearance  of  the  filtrate  caused  by 
minute,  resinous  drops.  Wash  flask  and  filter  once  with  10  cc. 
of  15  per  cent,  alcohol,  dry  the  filter  in  the  flask  and  weigh  both. 
Finally,  apply  a  correction  on  account  of  the  solubiHty  of 
santonin  in  the  alcohol  used.  Every  10  grams  of  filtrate  contain 
6  mg.  of  santonin.     Santonin  by  this  method  is  crystalline, 

^  Archiv  der  Pliarmacie  237,  251  (1899). 


284  DETECTION   OF   POISONS 

and  usually  faintly  yellow.  J.  Katz  found  the  quantity  of 
santonin  in  wormseed  to  vary  between  1.2 1  and  3.16  (average 
2.42)  per  cent.  This  method  is  based  upon  the  fact  that  the 
santonin  in  10  grams  of  wormseed  is  easily  soluble  in  50  cc.  of  hot 
15  per  cent,  alcohol,  whereas  only  a  very  little  resin  is  dissolved 
by  this  dilute  alcohol.  As  this  dilute,  alcoholic  solution  cools, 
santonin  crystallizes  out  almost  quantitatively. 

Troches  of  Santonin. — To  estimate  the  quantity  of  santonin  in  troches,  made 
from  this  substance  and  sugar,  directly  extract  the  finely  ground  mixture 
with  hot  chloroform.  The  santonin  can  usually  be  weighed  without  further 
purification. 

Chocolate  Troches  of  Santonin.^ — In  a  somewhat  simpler  form,  the  method 
described  above  may  be  used  to  estimate  santonin  in  chocolate  troches.  Weigh 
3  or  4  troches  and  boil  15  minutes  under  a  reflux  condenser  with  5  grams  of  barium 
hydroxide  and  100  cc.  of  water.  Saturate  the  liquid  when  cold  with  carbon 
dioxide.  Filter,  wash  the  residue  with  water  and  evaporate  the  brownish  filtrate 
to  100  cc.  Warm  the  liquid  and  add  10  cc.  of  dilute  hydrochloric  acid.  Three 
extractions  with  chloroform  yield  nearly  pure  santonin.  To  get  santonin  crys- 
tals almost  white  and  ready  for  weighing,  evaporate  the  chloroform  solution  and 
expel  the  last  traces  of  chloroform  by  adding  a  few  cc.  of  ether.  If  santonin  is 
impure  from  traces  of  fatty  acids,  boil  once  with  10  cc.  of  petroleum  ether  and 
filter  when  cold.     Santonin  is  nearly  insoluble  in  cold  petroleum  ether. 

Santonin  can  be  detected  and  estimated  in  toxicological  analysis  in  a  similar 
manner.  Acidify  the  material  with  hydrochloric  acid,  extract  with  chloroform 
and  treat  the  chloroform  residue  with  barium  hydroxide  solution  as  described 
above. 

Estimation  of  Solanine  in  Potatoes^ 

I.  O.  Schmiedeberg  and  G.  Meyer's  Method. — Mix  500 

grams  of  finely  grated  potatoes  with  water  and  press  out  the 
liquid.  Decant  the  liquid  from  the  deposit  of  starch.  Again 
mix  the  starch  with  water  and  decant  the  latter  when  the 
starch  has  settled.  Neutralize  the  entire  liquid  with  ammonia 
and  evaporate  to  the  consistency  of  an  extract.  In  the  mean- 
time mix  the  press  cake  with  several  times  its  volume  of  boiling 
alcohol.  Press  out  the  alcohol  completely  after  several  hours. 
Make  two  such  extractions.  Filter  the  combined  alcoholic 
extracts  and  wash  the  residue  (starch)  upon  the  filter  with 
alcohol.     The  aqueous  liquid  from  the  potatoes  contains  very 

^  See  page  217. 


QUANTITATIVE   ESTIMATION   01'   ALKALOIDS  285 

little  solanine.  To  isolate  this  small  quantity,  use  the  alcoholic 
filtrate  to  extract  the  residue  from  the  aqueous  extract  and 
again  filter.  Wash  the  insoluble  part  with  hot  alcohol.  The 
alcoholic  filtrate  after  half  an  hour  usually  deposits  some 
crystals  of  asparagine.^  Separate  the  supernatant  liquid  from 
these  crystals  and  evaporate  upon  the  water  bath  to  the  con- 
sistency of  an  extract.  Dissolve  the  residue  in  water  con- 
taining sulphuric  acid,  filter  and  wash.  Warm  the  clear 
liquid  very  gently,  saturate  with  ammonia  and  set  aside  for  a 
day.  Solanine  appears  in  small  crystals.  Collect  the  deposit 
upon  a  weighed  filter,  wash  first  with  water  and  then  with  ether, 
dry  at  ioo°  and  weigh. 

2.  F.  von  Morgenstem's^  Method. — Express  as  much 
liquid  as  possible  from  200  grams  of  finely  grated  potatoes  by 
by  means  of  a  press.  Make  two  separate  extractions  of  the 
press  cake  with  water  and  express  the  liquid  thoroughly  each 
time.  Precipitate  protein  sustances  from  the  combined  liquid 
by  adding  0.5  cc.  of  acetic  acid  and  warming  for  i  hour  upon  the 
water  bath.  Filter,  evaporate  the  filtrate  to  a  syrup,  stir  and 
add  gradually  hot  96  per  cent,  alcohol  until  cloudiness  ceases.^ 
Decant  the  solution  after  12  hours  and  extract  the  residue 
containing  sugars  and  dextrins  twice  with  hot  alcohol.  Evapo- 
rate the  combined  alcoholic  extracts  upon  the  water-bath,  warm 
the  residue  with  some  water  containing  acetic  acid  and  filter. 
Heat  the  filtrate  to  boiling  and  add  ammonia  drop  by  drop  to 
precipitate  solanine.  After  standing  for  5  minutes  upon  the 
water-bath  the  base  separates  in  flocks  that  are  easily  filtered. 
Wash  the  precipitate  with  water  containing  ammonia,  dissolve 
in  boiling  alcohol  and  treat  this  solution  as  follows.     Evaporate 

"■  Asparagine  is  the  amide  of  aspartic  acid,  or  mono-amino-succinic  acid, 
H2N.CH-COOH 

I  -|-  H2O.     It  appears  in  shinina;,  rhombic   crystals   that 

CH2-CO.NH2 
dissolve  rather  easily  in  hot  water  but  less  easily  in  alcohol  or  ether.     Lsevo- 
asparagine  is  widespread  in  the  plant  kingdom  in  seeds. 

^  Landwirtschaftliche  Versuchsstatibn  65,  301  (1907). 

^  To  extract  those  parts  of  the  potato  plant,  which  can  be  dried  at  100°  and 
reduced  to  a  fine  powder,  heat  to  boihng  several  times  with  water  containing 
acetic  acid  and  filter  each  time. 


286  DETECTION   OF   POISONS 

upon  the  water-bath  and  dissolve  the  residue  in  water  containing 
acetic  acid.  Filter,  heat  the  filtrate  to  boiling  and  precipitate 
solanine  with  ammonia.  Collect  the  pure  white  flocks  of  sola- 
nine  from  this  second  precipitation  upon  a  filter  that  has  been 
dried  at  90°  and  weighed.  Wash  with  2  per  cent,  ammonia  and 
dry  at  90°  to  constant  weight. 

Notes.! — V.  Moigenstern  obtained  on  the  average  by  this  method  0.0125  per 
cent,  of  solanine  in  table  potatoes  and  0.0058  per  cent,  in  those  used  as  forage. 
The  yield  of  solanine  from  yellow  tubers  upon  the  average  was  less  than  from  red. 
Tubers  grown  upon  sandy  soil  were  richer  in  solanine  than  were  those  from  humus 
soil.  Moisture  and  abundance  of  humus  appear  to  diminish  the  quantity  of 
solanine.  A  nitrogenous  fertilizer  increased  the  quantity  of  solanine,  a  potash 
fertilizer  lowered  it  and  a  phosphate  fertilizer  appeared  to  have  little  effect. 
There  was  less  solanine  in  large  than  in  small  potatoes  of  the  same  vaiiety. 
Solanine  first  appears  to  increase  during  the  process  of  germination.  Passing 
into  sprouts,  without  wholly  disappearing  from  the  tubeis,  solanine  increases 
with  the  growth  of  the  plant.  As  growth  advances  the  distribution  of  solanine 
in  the  different  parts  of  the  plant  is  indication  of  a  tendency  on  the  part  of  the 
plant  to  withdraw  solanine  from  the  older  sprouts  and  spread  it  throughout  the 
young  organs.  Consequently  solanine  may  serve  first  of  all  as  the  natural 
protector  of  the  plant,  especially  of  the  growing  parts. 

Estimation  of  Alkaloids  in  Nvx  Vomica 

(C.  C.  Keller') 

Remove  fat  from  nux  vomica  by  treating  15  grams  of  the 
well-dried  and  finely  powdered  drug  in  a  250  cc.  Erlenmeyer 
flask  two  or  three  times  with  30  cc.  portions  of  ether.  Shake 
thoroughly  for  5  minutes.  Pour  these  ether  washings  into  a 
flask,  and,  since  they  contain  a  little  alkaloid,  extract  dissolved 
alkaloid  with  5  cc.  of  o.i  n-hydrochloric  acid  and  10  cc.  of  water. 
Repeat  the  extraction  of  the  ether  layer,  separated  from  the 
aqueous  solution,  using  water  instead  of  acid.  Add  100  cc.  of 
ether,  50  grams  of  chloroform  and  10  grams  of  10  per  cent,  am- 
monium hydroxide  solution  to  the  powdered  nux  vomica  free 
from  fat.  Shake  thoroughly  for  30  minutes  and  add  to  this 
mixture  the  hydrochloric  acid  solution  used  in  extracting  alka- 

1  Festschrift  presented  at  the  fiftieth  anniversary  of  the  founding  of  the  Swiss 
Pharmaceutical  Association.  Abstract  in  Zeitschrift  fiir  analytische  Chemie, 
23,  491  (i^ 


QUANTITATIVE    ESTIMATION    OF    AI.KALOIUS  287 

loid  from  the  first  ether  washings.  Again  shake  thoroughly 
and,  when  the  hquids  have  separated  clear,  pour  loo  grams  of 
ether-chloroform  solution  through  a  small  filter  into  a  weighed 
Erlenmeyer  flask.  Distil  the  chloroform  and  ether  as  com- 
pletely as  possible.  The  alkaloids  usually  appear  as  colorless 
varnishes  which  persistently  retain  chloroform.  To  remove  the 
latter,  pour  a  few  cc.  of  absolute  alcohol  upon  the  residue  and 
expel  completely  upon  the  water-bath.  Repeat  this  treatment 
2-3  times.  This  will  give  crystalhne  alkaloids  which  can  be  dried 
at  100°  to  constant  weight. 

Method  of  the  German  Pharmacopoeia 

I.  In  Nux  Vomica. — Place  15  grams  of  nux  vomica,  ground  mediumly  fine 
and  dried  at  100°,  in  an  Erlenmeyer  flask  and  add  100  grams  of  ether  and  50 
grams  of  chloroform.  Shake  vigorously  and  add  10  cc.  of  a  mixture  of  2  parts 
of  sodium  hydroxide  solution  and  i  part  of  water.  Shake  at  frequent  intervals 
and  let  the  mixture  stand  foi  3  hours.  Then  add  15  cc.  more  of  watei,  cr  enough 
to  cause  the  powder  after  vigorous  shaking  to  gather  into  balls  and  leave  the  super- 
natant ethei-chloroform  solution  perfectly  clear.  After  i  hour  filter  100  grams 
of  the  clear  ether-chloroform  solution  through  a  dry  filter  kept  well  covered. 
Collect  the  filtrate  in  a  small  flask  and  distil  about  half  the  solvent.  Transfer 
the  residual  ether-chloroform  solution  to  a  separating  funnel,  rinse  the  flask 
3  times  with  5  cc.  portions  of  a  mixture  of  3  parts  of  ether  and  i  part  of  chloroform. 
Extract  the  combined  solvent  with  10  cc.  of  o.i  n-hydrochloric  acid.  Add  enough 
ether  to  cause  the  ether-chloroform  solution  to  rise  to  the  top  of  the  acid  liquid 
and  pass  the  latter  through  a  small  filter  moistened  with  water  into  a  100  cc. 
flask.  Then  extract  the  ether-chloroform  solution  with  3  additional  10  cc.  por- 
tions of  water.  Pass  these  extracts  through  the  same  filter,  wash  the  latter  with 
water  and  dilute  the  total  liquid  to  100  cc.  Finally  measure  50  cc.  of  this  solution 
into  a  flask  holding  about  200  cc,  add  about  50  cc.  of  water  and  sufiicient  ether 
to  make  a  layer  i  cm.  deep.  Add  5  drops  of  iodeosine  solution  and  run  in  enough 
o.oi  n-potassium  hydroxide  solution,  shaking  vigorously  after  each  addition,  to 
turn  the  aqueous  layei  a  permanent  pale  red. 

Calculation. — 100  grams  (=  alkaloids  from  10  grams  of  nux  vomica)  of  the 
original  150  grams  of  ether-chloroform  mixture  were  used.  The  alkaloids  were 
dissolved  by  10  cc.  of  o.i  n-hydrochloiic  acid  and  the  volume  was  brought  to 
ICO  cc.  The  excess  of  acid  in  50  cc.  of  this  solution  ( =  5  grams  of  nux  vomica) 
was  determined  b}'  titration  with  o.or  n-potassium  hvdroxide.  If  strychnine 
and  brucine  are  present  in  nux  vomica  in  equal  amount,  the  average  equivalent 
weight  of  the  two  alkaloids  is  364.  Therefore  1000  cc.  of  0.1  n-hydrochloric 
acid  correspond  to  36.4  grams  of  alkaloids. 

Example.' — Suppose  that  the  titration  of  the  excess  of  acid  in  50  cc.  of  solution 
required  15.6  cc.  of  o.or  n-potassium  hydroxide  =  1.56  cc.  of  o.r  n-alkali.     Then 


288  DETECTION   OF  POISONS 

5  —  1.56  =  3.44  cc.  of  0.1  n-hydrochloric  acid  are  combined  with  the  alkaloids 
in  s  grams  of  nux  vomica.     According  to  the  proportion 

Cc.  0.1  n-HCl:  Grams  alkaloid  • 

1000  :  36.4  =  3.44  :  X     (x  =  0.12522) 

3.44  cc.  of  0.1  n-acid  are  combined  with  0.12522  gram  of  alkaloid,  correspond- 
ing to  an  alkaloid  content  of  20  X  0.12522  =  2.50  per  cent.  The  German 
Pharmacopoeia  places  this  percentage  as  the  minimum  for  total  alkaloids  in 
nux  vomica. 

2.  In  Extract  of  Nixx  Vomica. — Dissolve  i  gram  of  extract  in  an  Erlenmeyer 
flask  in  5  grams  of  water  and  5  grams  of  absolute  alcohol,  and  add  50  grams  of 
ether  and  20  grams  of  chloroform  to  this  solution.  Shake  vigorously  and  add  10 
cc.  of  sodium  carbonate  solution  (i  .-3).  Let  the  mixture  stand  and  agitate  at 
frequent  intervals  for  an  hour.  *  Then  pass  50  grams  of  the  clear  ether-chloroform 
solution  through  a  dry  filter  kept  well  covered,  and  receive  the  filtrate  in  a  flask. 
Distil  half  the  solvent  and  pour  the  remainder  into  a  separating  funnel.  Wash 
the  flask  three  times  with  5  cc.  portions  of  a  mixture  of  3  parts  of  ether  and 
I  part  of  chloroform.  Thoroughly  extract  the  total  ether-chloroform  solution 
with  so  cc.  of  o.oi  n-hydrochloric  acid.  When  the  liquids  have  separated 
clear,  if  necessary,  after  addition  of  enough  ether  to  bring  the  ether-chloro- 
form solution  to  the  surface,  pass  the  acid  solution  through  a  small  filter 
moistened  with  water  and  receive  the  filtrate  in  a  200  cc.  flask.  Make  three 
extractions  of  the  ether-chloroform  solution  with  10  cc.  portions  of  water,  and 
pass  these  washings  through  the  same  filter.  Finally,  wash  the  filter  with  water 
and  bring  the  entire  solution  to  100  cc.  Add  enough  ether  to  make  a  layer  i  cm. 
thick  and  5  drops  of  iodeosine  solution.  Run  in  o.oi  n-potassium  hydroxide 
solution,  shaking  vigorously  after  each  addition,  until  the  aqueous  solution  is 
pale  red. 

Calculation. — Only  50  grams,  or  two-thirds  of  the  original  75  grams  of  alcohol- 
ether-chloroform  mixture,  were  used.  The  alkaloids  in  0.666  gram  of  nux 
vomica  extract  were  in  this  volume  of  solvent.  Alkaloids  were  dissolved  in  50 
cc.  of  O.OI  n-hydrochloric  acid  and  excess  of  acid  determined  by  titration  with 
O.OI  n-potassium  hydroxide  solution. 

Example. — Suppose  that  18  cc.  of  o.oi  n-potassium  hydroxide  solution  were 
used  in  this  titration.  Then  50  —  18  =  32  cc.  of  o.oi  n-acid  were  combined 
with  the  alkaloids  in  0.666  gram    of   extract.     According  to  the  proportion 

Cc.  o.oi  n-HCl  :  Grams  alkaloids 

1000  :  3.64  =  32  :  X     (x  =  0.11648) 

0.666  gram  of  extract  contains  0.11648  gram  of  alkaloids,  corresponding  to  17.47 
per  cent.  The  German  Pharmacopoeia  places  this  percentage  as  the  minimum 
for  total  alkaloids  in  extract  of  nux  vomica. 

3.  In  Tincture  of  Nux  Vomica. — Evaporate  50  grams  of  tincture  of  nux  vomica 
in  a  weighed  dish  to  10  grams.  Wash  this  residue  into  an  Erlenmeyer  flask  and 
rinse  with  5  grams  of  absolute  alcohol.  Add  50  grams  of  ether  and  20  grams  of 
chloroform  and  shake  vigorously.  Then  add  10  cc.  of  sodium  carbonate  solution 
(i  :  3)  which  has  been  previously  employed  in  rinsing  the  dish  used  in  evaporating 
the   tincture.     Let   this   mixture   stand   an   hour,  shaking  vigourously  at  fre- 


QUANTITATIVE   ESTIMATION   OF   ALKALOIDS  289 

quent  intervals.  Filter  50  grams  of  the  clear  ether-chloroform  solution.  To  ex- 
tract alkaloids,  use  40  cc.  of  o.oi  n-hydrochloric  acid.  In  other  respects,  the 
esti'mation  of  alkaloids  is  the  same  as  described  for  extiact  of  nux  vomica. 

Calculation. — This  is  the  same  as  that  given  for  extract  of  nux  vomica.  Only 
two-thirds  (=  2>2,-2>  grams)  of  the  original  weight  of  nux  vomica  tincture  were 
used.  The  alkaloids  were  dissolved  in  40  cc.  of  o.oi  n-hydrochloric  acid  and  ex- 
cess of  acid  was  determined  by  titration  with  o.oi  n-potassium  hydroxide 
solution.  If  17  cc.  of  the  latter  solution  were  used,  then  40  —  17  =  23  cc.  of 
O.OI  n-hydrochloric  acid  were  combined  with  the  alkaloids  in  2>Z-Z  grams  of  the 
tincture.     According  to  the  proportion 

Cc.  O.OI  n-HCl  :  Grams  alkaloids 

1000  :  3.64  =  23  :  x  (x  =  0.08372) 

this  weight  of  tincture  contains  0.08372  gram  of  alkaloids,  corresponding  to  2.51 
per  cent.  The  German  Pharmacopoeia  places  this  percentage  as  the  minimum 
for  total  alkaloids  in  tincture  of  nux  vomica.  Brucine  and  strychnine  are  assumed 
to  be  present  in  equal  quantity. 

Estimation  of  Alkaloids  in  Nux  Vomica  and  Its  Preparations  by  Means 

of  Picrolonic  Acid 

(H.  Matthes  and  O.  Rammstedt) 

Nux  vomica  upon  the  average  contains  strychnine  and 
brucine  in  equal  quantity  combined  with  tannic  acid.  Alkaline 
hydroxide  or  carbonate  solutions  Hberate  the  alkaloids  from 
their  salts.  The  free  bases  are  then  extracted  with  an  ether- 
chloroform  mixture.  The  solvent  is  reduced  to  smaller  volume 
by  evaporation  or  distillation  and  the  alkaloids  are  precipitated 
with  picrolonic  acid. 

Strychnine  picrolonate,  C21H22N2O2.C10H8N4O5  (Mol.  Wt. 
598)  melts  with  decomposition  at  286°. 

Brucine  picrolonate,  C23H26N2O4.C10H8N4O5  (AIol.  Wt.  658) 
melts  with  decomposition  at  277°. 

I.  Extract  of  Nux  Vomica. — Dissolve  i  gram  of  extract  in  5 
grams  of  absolute  alcohol  and  5  grams  of  water.  Shake  well 
with  50  grams  of  ether  and  20  grams  of  chloroform.  Add  10  cc 
of  sodium  carbonate  solution  (i  :  2)  and  again  shake  thoroughly 
for  10  minutes.  Let  the  mixture  stand  at  rest  for  20  minutes. 
Pass  50  grams  of  the  ether-chloroform  mixture  through  a  dry, 
double,  creased  filter  and  evaporate  half  the  solvent  in  a  beaker. 
Add  about  5  cc.  of  o.  i  n-alcoholic  picrolonic  acid  to  the  warm 

19 


290  DETECTION   OP   POISONS 

solution.  A  yellow  crystalline  precipitate  of  strychnine  and 
brucine  picrolonates  soon  appears.  After  24  hours  collect  the 
mixed  picrolonates  in  a  weighed  Gooch  crucible.  Wash 
excess  of  picrolonic  acid  from  the  precipitate  with  2  cc.  of  an 
alcohol-ether  mixture  (i  :3),  dry  30  minutes  at  110°,  cool  in 
desiccator  and  weigh. 

Calculation. — -Use  the  mean  molecular  weight  of  brucine  and  strychnine 
picrolonates (  =  628)  and  also  the  mean  molecular  weight  of  brucine  and  strych- 
nine (  =  364).     The  proportion  is 

Grams  picrolonate   Grams  strychnine         Wt.  of  pre- 
mixture  :      and  brucine         =       cipitate :  x. 

628  :  364 

Since  the  quotient  364  :628  =  0.5798,  the  weight  of  mixed  alkaloids  is  obtained 
by  multiplying  the  weight  of  the  picrolonate  precipitate  by  this  quantity.  This 
precipitate  represents  total  alkaloids  in  0.666  gram  of  extract  of  nux  vomica, 
for  only  two- thirds  (=50  grams)  of  the  original  75  grams  (5  grams  of  alcohol  + 
50  grams  of  ether  +  20  grams  of  chloroform)  of  solvent,  containing  the  alkaloids 
in  I  gram  of  extract  of  nux  vomica,  were  used. 

2.  Tincture  of  Nux  Vomica. — Evaporate  50  grams  of  tincture 
in  an  Erlenmeyer  flask  to  10  cc.  Cool  and  shake  well  with  5  cc. 
of  absolute  alcohol,  50  grams  of  ether  and  20  grams  of  chloro- 
form. Add  10  cc.  of  sodium  carbonate  solution  (i  :  2)  and  shake 
again  for  10  minutes.  After  20  minutes  pass  50  grams 
(=  two-thirds  of  the  original  mixture)  of  the  ether-chloroform 
mixture  through  a  double,  creased  filter.  Evaporate  half  the 
solvent  in  a  beaker  and  add  5  cc.  of  o.i  n-alcoholic  picrolonic 
acid  to  the  warm  residue.  Treat  the  picrolonate  precipitate  as 
described  above. 

3.  Nux  Vomica. — Exhaust  15  grams  of  powdered  nux  vomica, 
previously  dried  at  100,  by  thoroughly  agitating  with  100  grams 
of  ether  and  50  grams  of  chloroform.  Then  add  10  cc.  of  a 
mixture  of  2  parts  of  15  per  cent,  sodium  hydroxide  solution  and 
I  part  of  water  and  shake  again  for  10  minutes.  Add  an  ad- 
ditional 15  cc.  of  water,  or  enough  to  cause  the  powder  to  gather 
into  balls  after  vigorous  agitation  and  leave  the  supernatant 
ether-chloroform  mixture  clear.  After  30  minutes  pass  the 
clear  ether-chloroform  solution  through  a  dry,  double,  creased 
filter.     Evaporate  50  cc.  of  the  filtrate  in  a  beaker  nearly  to 


QUANTITATIVE   ESTIMATION    OF   ALKALOIDS  291 

dryness  and  add  a  second  50  cc.  portion  of  filtrate  to  the 
residue,  bringing  everything  into  solution  (  =  alkaloids  from  10 
grams  of  nux  vomica).  Add  5  cc.  of  o.i  n-alcoholic  picrolonic 
acid  and  treat  the  precipitate  as  previously  described. 

Estimation  of  Strychnine  in  Mixtures  of  Nux  Vomica  Alkaloids 

(Gordin'si  Modification  of  Keller's  Method) 

Strong  nitric  acid,  gently  heated  with  a  solution  of  strych- 
nine and  brucine  in  3  per  cent,  sulphuric  acid,  is  without  action 
upon  the  former  alkaloid.  But  brucine  is  converted  into  non- 
basic  substances  not  extracted  by  chloroform  from  an  alkaline 
solution. 

Procedure.^ — Dissolve  the  mixed  alkaloids  (0.2  —  0.3  gram) 
upon  the  water-bath  in  15  cc.  of  3  per  cent,  sulphuric  acid  and 
add  3  cc.  of  a  diluted  nitric  acid  (equal  parts  of  68-69  P^r  cent, 
acid  (sp.  gr.  1.42)  and  water)  to  the  cold  solution.  Pour  the 
mixture  into  a  separating  funnel  after  exactly  10  minutes  and 
add  sodium  hydroxide  solution  in  excess.  Extract  strychnine 
3-4  times  with  chloroform.  Pass  the  chloroform  solution 
through  a  double  filter  into  a  small  weighed  flask,  wash  the 
filter  with  a  little  chloroform,  add  2  cc.  of  pure  amyl  alcohol  and 
distil  to  dryness  upon  the  water-bath.  Remove  the  last  traces 
of  liquid,  consisting  mainly  of  amyl  alcohol,  by  forcing  a  current 
of  air  through  the  flask  warmed  upon  the  water-bath.  Finally 
dry  the  residue  2  hours  at  135-140°  and  weigh  when  cold.  In 
this  way  a  very  pure,  white  strychnine  free  from  brucine  is 
obtained. 

Notes. — According  to  Gordin,  ammonia  cannot  be  substituted  for  sodium 
hydroxide,  for  it  gives  colored  strychnine.  Amyl  alcohol  is  added  to  the  chloro- 
form solution  to  prevent  strychnine  crj^stals  from  being  carried  by  decrepitation 
into  the  condenser  during  distillation. 

Estimation  of  Theobromine  and  Caffeine  in  Cacao  and  Chocolate = 

Cacao  and  its  preparations  contain  only  very  Kttle  cafi'eine 
which  is  usually  determined  with  theobromine. 

^  Archiv  der  Pharmazie  240,  643  (1902). 
H.  Beckurts,  Archiv  der  Pharmazie,  244,  486    (1906). 


292  DETECTION   OP   POISONS 

Boil  6  grams  of  powdered  cacao,  or  12  grams  of  chocolate,  for 
30  minutes  under  a  reflux  condenser  in  a  weighed  liter  flask  with 
200  grams  of  a  mixture  of  197  grams  of  water  and  3  grams  of 
dilute  sulphuric  acid.  Then  add  400  grams  of  water  and  8 
grams  of  finely  powdered  magnesia  and  boil  for  an  hour  longer. 
When  the  mixture  is  cold,  add  exactly  enough  water  by  weight 
to  replace  what  has  been  lost  by  evaporation.  After  the  mix- 
ture has  settled,  filter  500  grams  of  solution  (=  5  grams  of  cacao 
or  10  grams  of  chocolate)  and  evaporate  the  filtrate  to  dryness 
in  a  dish  either  by  itself  or  with  some  quartz  sand.  Triturate 
the  residue  and  exhaust  in  a  Soxhlet  tube  with  chloroform. 

In  case  of  evaporation  without  quartz  sand,  rub  the  residue 
with  a  few  drops  of  water,  transfer  to  a  separating  funnel  with 
10  cc.  of  water  and  extract  8  times  with  50  cc.  portions  of  hot 
chloroform.  Pass  the  chloroform  extract  through  a  dry  filter 
into  a  tared  flask,  distil  the  chloroform  and  dry  the  residue 
(=  theobromine  and  caffeine)  at  100°  to  constant  weight. 

Carbon  tetrachloride  is  used  to  separate  theobromine  from 
caffeine,  the  latter  alkaloid  alone  being  soluble  at  room  tem- 
perature. Let  the  weighed  residue  from  chloroform  stand  for  i 
hour  with  100  grams  of  carbon  tetrachloride  at  room  tem- 
perature. Shake  occasionally  and  then  filter.  Distil  carbon 
tetrachloride  and  extract  the  residue  repeatedly  with  water. 
Evaporate  the  aqueous  solution  in  a  weighed  dish  and  dry  the 
residue  (=  caffeine)  at  100°  to  constant  weight. 

Repeatedly  extract  the  theobromine,  insoluble  in  carbon  tetra- 
chloride, and  also  the  filter  paper  with  water.  Filter,  evaporate 
the  total  filtrate  and  weigh  the  residue  (  =  theobromine)  dried 
at  100°. 

Notes. — In  the  method  described  above,  H.  Beckurts  and  Fromme  eUminate 
injurious  effects  due  to  concentration  by  boiling  with  dilute  sulphuric  acid. 
Xanthine  bases  are  set  free  from  combination  with  organic  acid  and  recombined 
with  sulphuric  acid.  Magnesia  sets  these  bases  free  from  their  sulphates  and 
at  the  same  time  holds  back  coloring  matter  and  fat,  thus  eliminating  these 
impurities. 

Theobromine,  3,7-dimethyl-xanthine,  C7H8N4O2,  is  a  white  powder  consisting 
of  microscopic  needles  having  a  bitter  taste.  It  dissolves  in  3282  parts  of  cold 
and  148  parts  of  boiling  water;  in  422  parts  of  boiling  absolute  alcohol;  and  in 


QUANTITATIVE   ESTIMATION    OF   ALKALOIDS  293 

105  parts  of  boiling  chloroform.  Theobromine  solutions  are  neutral.  This 
alkaloid  acts  both  as  an  acid  and  as  a  base  and  therefore  is  soluble  in  both  acid 
and  alkahne  solutions.  The  salts  with  acids  crystallize  well  but  are  not  very 
stable.  These  theobromine  salts  are  partially  decomposed  into  theobromine 
and  acid  in  presence  of  much  water,  or,  if  the  given  acid  is  volatile,  by  heating  at 
100°.  Theobromine  is  isomeric  with  theophylline,  or  1,3-dimethyl-xanthine, 
and  paraxanthine,  or  1,7-dimethyl-xan thine: 

HN— CO  (i)CH3.N— CO  (i)CH3.N— CO 

I       I         /CH3(7)  I       I         H  II         yClUy) 

OC    C— N<  OC     C— N\  OC     C— N< 

I       II        >CH  I      II        >CH  I      II        >CH 

(3)CH3.N— C— N^  (3)CH3.N— C— N^  HN=C— N^ 

Theobromine  Theophylline  Paraxanthine 

Theophylline  occurs  in  tea  leaves  and  paraxanthine  has  been  isolated  from  human 
urine.     The  latter  is  therefore  called  urotheobromine. 

Estimation  of  Alkaloids  in  Leaves  of  Atropa  Belladonna,  Hyoscyamus 
Niger  and  Datura  Strammonium 

(E.  Schmidt's  Modification  of  Keller's  Method^) 

Shake  vigorously  10  grams  of  finely  powdered  leaves,  dried  to  constant  weight 
over  quicklime,  in  an  Erlenmeyer  flask  with  90  grams  of  ether  and  30  grams  of 
chloroform.  Add  10  cc.  of  10  per  cent  sodium  hydroxide  solution  and  shake 
vigorously  and  often  for  3  hours.  Then  add  10  cc.  of  water,  or  enough  to  cause 
the  powder  to  gather  into  balls  when  thoroughly  shaken.  After  i  hour  pass  60 
grams  of  the  ether-chloroform  extract  (=  5  grams  of  leaves)  through  a  dry 
filter  kept  well  covered.  Distil  60  cc.  of  this  filtrate  to  half  its  voluine  to  remove 
ammonia,  and  transfer  the  deep  green  solution  to  a  separating  funnel,  rinsing 
the  flask  with  three  5  cc.  portions  of  ether.  Shake  the  combined  extracts  well 
with  10  cc.  of  o.oi  n-hydrochloric  acid.  Add  enough  ether  to  cause  the  ether- 
chloroform  solution  to  rise  to  the  top,  and  pass  the  acid  solution  through  a  moist 
filter  into  a  200  cc.  glass  stoppered  flask.  Shake  the  ether-chloroform  solution 
3  times  with  10  cc.  portions  of  water,  pouring  these  extracts  through  the  same 
filter  and  washing  the  latter  with  enough  water  to  bring  the  total  volume  to  100 
cc.  Add  enough  ether  to  make  a  layer  i  cm.  deep  and  5  drops  of  iodeosine  solu- 
tion. Having  determined  beforehand  the  exact  relation  of  acid  to  alkali,  titrate 
excess  of  o.oi  n-hydrochloric  acid  with  o.oi  n-potassium  hj'droxide  solution. 
The  calculation  is  the  same  as  that  for  extract  of  belladonna  (see  page  294). 

Notes. — Using  this  method,  E.  Schmidt  obtained  0.4  per  cent,  of  alkaloid  in 
wild  belladonna  leaves  but  only  0.26  per  cent,  in  cultivated  leaves.  The  average 
of  many  determinations  gave  0.4  per  cent,  in  strammonium  leaves  and  0.27-0.28 
per  cent,  in  hyoscyamus  leaves  without  stalks.  Alkaloids  were  calculated  as 
atropine. 

Sodium  hydroxide  solution  liberates  alkaloids  from  the  acids  with  which  they 
are  naturally  combined  in  the  plant,  for  example: 

(Ci7H23N03)2.H2S042  +  2NaOH  =  2C17H23NO3  +  2H2O  +  Xa2S04. 

1  Apotheker-Zeitung  15,  13. 

^  The  formula  of  atropine  sulphate  used  in  medicine. 


294  DETECTION   OP   POISONS 

Estimation  of  Alkaloids  in  Extract  of  Belladonna 

(German  Pharmacopoeia) 

Dissolve  2  grams  of  extract  cf  belladonna  in  an  Erlenmeyer  flask  in  5  grams  of 
water  and  5  grams  of  absolute  alcohol,  and  add  50  grams  of  ether  and  20  grams  of 
chloroform  to  this  solution.  Shake  vigorously  and  add  10  cc.  of  sodium  carbonate 
solution  (i  :3).  Let  the  mixture  stand  and  agitate  at  frequent  intervals  for  an 
hour.  Then  pass  50  grams  of  the  clear  ether-chloroform  solution  through  a  dry 
filter  kept  well  covered,  and  receive  the  filtrate  in  a  flask.  Distil  half  the  solvent 
and  pour  the  remainder  into  a  separating  funnel.  Wash  the  flask  three  times 
with  5  cc.  portions  of  ether.  Thoroughly  extract  the  total  ether-chloroform  so- 
lution with  20  cc.  of  o.oi  n-hydrochloric  acid.  When  the  litfuids  have  separated 
clear,  if  necessary,  after  addition  of  enough  ether  to  bring  the  ether-chloro- 
form solution  to  the  surface,  pass  the  acid  solution  through  a  small  filter  moist- 
ened with  water  and  receive  the  filtrate  in  a  200  cc.  flask,  ^ake  three  extrac- 
tions of  the  ether-chloroform  solution  with  10  cc.  portions  of  water,  and  pass  these 
washings  through  the  same  filter.  Finally,  wash  the  filter  with  water  and  bring 
the  entire  solution  to  100  cc.  Add  enough  ether  to  make  a  layer  i  cm.  thick 
and  5  drops  of  iodeosine  solution.  Ruh  in  o.oi  n-potassium  hydroxide  solution, 
shaking  vigorously  after  each  addition,  until  the  aqueous  solution  is  pale  red. 

Calculation. — Sodium  carbonate  like  sodium  hydroxide  liberates  the  alkaloids 
atropine  and  hyoscyamine,  from  their  salts  in  belladonna  leaves: 

(Ci7H23N03)2.H2S04  +  NaaCOg  =  2C17H23NO3  +  Na2S04  +  CO2  +  H2O. 
The  free  alkaloids  dissolve  in  the  alcohol-ether-chloroform  mixture.  Fifty  grams 
of  this  solution  ( =  alkaloids  from  1.33  grams  of  extract)  are  extracted  with  20  cc. 
of  O.OI  n-hydrochloric  acid,  the  alkaloids  passing  into  the  aqueous  solution  as 
salts  of  hydrochloric  acid  (C17H23NO3.HCI).  Excess  of  acid  in  this  solution  is 
determined  by  titration.  If  13  cc.  of  o.oi  n-potassium  hydroxide  solution  are 
used,  then  20  -  13  =  7  cc.  of  o.oi  n-hydrochloric  acid  correspond  to  the  alkaloids 
in  1.33  grams  of  extract.  The  equivalent  weight  of  the  two  isomeric  bases, 
atropine  and  hyoscyamine,  being  289,  1000  cc.  of  o.oi  n-hydrochloric  acid  corre- 
spond to  2. 89  grams  of  alkaloids.     The  proportion 

1000  :  2.89  =  7  :  X     (x  =  0.02023) 

shows  that  1.33  grams  of  belladonna  extract  contain  0.02023  gram  of  alkaloids 
corresponding  to  1.51  per  cent.  The  German  Pharmacopoeia  places  this  per- 
centage as  the  minimum  for  total  alkaloids  in  extract  of  belladonna. 

Extract  of  Hyoscyamus 

The  alkaloids  in  2  grams  of  this  extract  are  determined  in  the  manner  described 
for  extract  of  belladonna.  Use  10  cc.  of  o.i  n-hydrochloric  acid  instead  of  20  cc. 
to  extract  alkaloids.  The  German  Pharmacopoeia  requires  that  not  more  than 
6.5  cc.  of  O.OI  n-potassium  hydroxide  solution  shall  be  used  in  titrating  the  excess 
of  hydrochloric  acid.  Therefore  10  —  6.5  =  3-5  cc.  of  o.oi  n-hydrochloric  acid 
are  combined  with  the  alkaloids  in  1.3  grams  of  henbane  extract  (=  two- thirds 
of  the  original  extract).     The  proportion 

1000:2.89  =  3-5 -x     (x  =  o.oioii) 


QUANTITATIVE    ESTIMATION    OF    ALKALOIDS  295 

shows  that  1.33  grams  of  extract  contain  o.oioi  gram  of  alkaloid,  corresponding 
to  0.76  per  cent.  This  percentage  is  placed  as  the  minimum  for  total  alicaloids  in 
henbane  extract. 

Assaying  OflBcinal  Extracts 

(K.  Merck') 

With  a  view  to  obviating  as  many  sources  of  error  as  possible,  K.  Merck  has 
proposed  the  following  procedures: 

Extract  of  Belladonna. — Dissolve  4  grains  of  extract  in  6  cc.  of  water  and  wash 
the  solution  into  a  separating  funnel  with  an  additional  10  cc.  Add  100  cc.  of 
ether,  shaking  well,  then  10  cc.  of  sodium  carbonate  solution  (i  -.t,)  and  shake  at 
once  for  5  minutes.  Stopper  the  funnel  and  let  the  mixture  stand  for  20  minutes. 
Then  pass  the  ether  layer  through  a  dry  filter  (10  cm.  in  diameter)  into  a  glass- 
stoppered  flask.  To  lessen  evaporation  of  ether  as  much  as  possible,  cover  the 
funnel  with  a  glass  plate.  If  an  emulsion  keeps  the  ether  from  separating  well, 
add  a  few  grams  of  powdered  tragacanth  at  the  end  of  the  time  stated  above. 
Shake  until  the  tragacanth  gathers  into  balls  in  the  aqueous  layer.  After  15 
minutes  decant  75  cc.  of  the  ether  layer.  To  check  results  by  making  more  than 
one  assay,  use  25  cc.  of  the  ether  solution  (=  i  gram  of  extract).  Test  a  clean 
glass-stoppered  flask  to  make  sure  that  it  does  not  give  up  alkali  to  the  water. 
Then  introduce  into  such  a  flask  50-60  cc.  of  water,  5  drops  of  iodeosine  solution 
and  20  cc.  of  ether.  Shake  and  add  0.0 1  n-hydrochloric  acid  until  the  aqueous 
layer  just  becomes  colorless  upon  shaking.  This  procedure  obviates  a  special 
determination  of  the  alkalinity  of  the  water,  since  the  resulting  mixture  is  brought 
to  the  neutral  point.  Now  add  25  cc.  of  the  ether  solution  of  the  alkaloid  and 
titrate  until  there  is  no  color.  Multiply  the  number  of  cc.  of  o.oi  n-hydrochloric 
acid  used  by  0.00289.^  The  product  is  the  quantity  of  alkaloid,  calculated  as 
atropine,  in  i  gram  of  belladonna  extract.  Upon  the  average  this  preparation 
contains  1.8  per  cent,  of  alkaloid. 

Extract  of  Cinchona. — Haematoxylin  is  frequently  an  unsatisfactory  indicator 
in  the  titration  of  cinchona  alkaloids,  because  the  color  change  is  slow  enough 
to  make  it  difficult  to  fix  the  end  point  exactly.  Therefore  E.  Merck  makes  a 
gravimetric  and  volumetric  determination  at  the  same  time  by  the  follo^-ing 
method: 

Dissolve  3  grams  of  aqueous  cinchona  extract  in  10  cc.  of  water  in  a  porcelain 
dish.  Pour  the  solution  into  a  250  cc.  shaking  flask,  rinsing  it  in  with  10  cc. 
of  water.  Add  150  cc.  of  ether  and  10  cc.  of  sodium  carbonate  solution  (1:3) 
to  this  mixture  and  shake  vigorously  for  10  minutes.  Cork  the  flask  and  let  the 
mixture  stand  at  rest  for  30  minutes.  This  extract  frequently  forms  an  emulsion. 
In  that  case  add  a  few  grams  of  tragacanth  powder  which  has  no  effect  upon  the 
result.  Pour  the  ether  solution  of  cinchona  alkaloids  as  rapidly  as  possible 
through  a  dry  creased  filter.     Use  50  cc.  (=  i  gram  of  cinchona  extract)  for  each 

1  Zeitschrift  fiir  analytische  Chemie  41,  584  (1902)  and  also  Merck's  Bericht 
iiber  das  Jahr  1900. 

2  289  =  the  equivalent  weight  of  the  two  isomeric  bases,  atropine  and  hyoscy- 
amine,  CnHasNOs. 


296  DETECTION   OF  POISONS 

determination.  Distil  the  solvent  from  the  50  cc.  in  a  weighed  100  cc.  flask  and 
dry  the  residue  in  an  air  bath  at  100-110°  to  constant  weight.  The  alkaloids 
obtained  are  nearly  colorless  or  faintly  yellow.  Having  ascertained  the  weight 
of  the  alkaloids,  proceed  with  the  titration.  Dissolve  the  residue  in  the  flask  in 
10  cc.  of  alcohol,  adding  50  cc.  of  water,  which  partially  precipitates  the  alkaloids, 
and  then  alcoholic  hsematoxylin  solution.^  Run  in  o.i  n-hydrochloric  acid  until 
the  alkaloids  again  dissolve  and  the  red  color  of  the  solution  passes  through 
reddish  yellow  into  a  pure  yellow.  The  mean  equivalent  weight  of  the  cinchona 
alkaloids  is  309.  Therefore  i  cc.  of  o.i  n-hydrochloric  acid  =  0.0309  gram  of 
alkaloid.  Upon  the  average,  officinal  aqueous  extract  of  cinchona  contains  9 
per  cent,  of  alkaloid. 

Extract  of  Ntix  Vomica. — Dissolve  0.1  gram  of  this  extract  in  a  flask  in  5 
grams  of  absolute  alcohol  and  10  grams  of  water.  Add  95  grams  of  ether  and 
shake  well.  Then  add  10  cc.  of  sodium  carbonate  solution  (1:3)  and  shake 
vigorously  at  once  for  about  10  minutes.  After  15  minutes  pour  the  ether  solu- 
tion as  rapidly  as  possible  through  a  creased  filter.  Weigh  in  a  flask  50  grams  of 
this  solution  (=  0.05  gram  of  the  original  extract),  having  previously  placed  in 
this  flask  a  neutral  mixture  of  50  cc.  of  water,  20  cc.  of  ether  and  5  drops  of  iod- 
eosine  solution.  Add  20  cc.  of  o.oi  n-hydrochloric  acid  and  titrate  with  o.oi 
n-potassium  hydroxide  solution  until  the  aqueous  layer  is  just  red. 

Calculation. — Since  strychnine  and  brucine  are  present  in  nux  vomica  in  nearly 
equal  parts,  the  mean  equivalent  weight  of  such  a  mixture  of  bases  (334  +  394)  :  2 
=  364.  Hence  0.00364  gram  of  the  mixed  alkaloids  neutralizes  i  cc.  of  o.oi 
n-hydrochloric  acid.  The  officinal  extract  of  nux  vomina  contains  18  per  cent, 
of  alkaloid. 

^  E.  Merck  advises  keeping  on  hand  an  alcoholic  solution  of  hsematoxylin, 
because  a  freshly  prepared  solution  usually  gives  a  blue-violet  instead  of  a  red 
color  change. 


CHAPTER  VII 

DETECTION  OF  CARBON  MONOXIDE  BLOOD,  BLOOD  STAINS 
AND  HUMAN  BLOOD 

I.  Carbon  Monoxide  Blood 

Carbon  monoxide  (CO)  has  a  direct  toxic  action  upon  the 
blood.  This  gas  passed  into  blood  displaces  loosely  bound 
oxygen  from  oxyhcemoglobin  forming  the  more  stable  carboxy- 
hasmoglobin.  The  latter  compound  is  cherry  red,  not  dichroic 
and  entirely  resistant  to  putrefaction  if  air  is  excluded.  In 
carbon  monoxide  poisoning  the  cherry  red  color  of  the  blood 
is  usually  noticed  at  once. 

Detection  of  Carbon  Monoxide  Blood 

1.  Boiling  Test. — Blood  containing  carbon  monoxide  gives  a 
brick  red  coagulum,  if  boiled  or  warmed  upon  the  water-bath. 
Ordinary  blood  gives  a  grayish  brown  or  brownish  black 
precipitate. 

2.  Sodixtm  Hydroxide  Test. — Carbon  monoxide  blood  shaken 
with  1-2  volumes  of  sodium  hydroxide  solution  (sp.  gr.  1.3  =  26.8 
per  cent.)  remains  red  and  in  a  thin  layer  is  the  color  of  red  lead 
or  vermilion.  Normal  blood  similarly  treated  is  almost  black 
and  in  a  thin  coating  upon  a  porcelain  plate  is  dark  greenish 
brown.  A  procedure  recommended  consists  in  diluting  the 
blood  with  6-10  times  its  volume  of  water  and  using  about  5 
drops  of  sodium  hydroxide  solution  to  10  cc.  of  diluted  blood. 
Even  gentle  warming  with  sodium  hydroxide  solution  Cio  per 
cent.  NaOH)  does  not  alter  the  red  color  of  this  carboxyhaemo- 
globin  solution,  whereas  a  solution  of  normal  human  blood 
becomes  greenish  to  dark  brown. 

3.  Basic  Lead  Acetate  Test. — Mix  4-5  volumes  of  basic 
lead  acetate  solution  in  a  test-tube  with  diluted  or  undiluted 
carbon  monoxide  blood  and  shake  vigorously  for  a  minute. 

297 


298  DETECTION   OF   POISONS 

Such  blood  remains  bright  red  but  normal  blood  is  first  brown- 
ish and  then  chocolate  to  greenish  brown. 

4.  Potassium  Ferrocyanide  Test. — Mix  undiluted  blood  (15 
cc.)  with  an  equal  volume  of  20  per  cent,  potassium  ferrocyanide 
solution  and  2  cc.  of  diluted  acetic  acid.^  Shake  the  mixture 
gently  and  a  coagulum  will  gradually  form.  That  from  normal 
blood  is  dark  brown  but  from  blood  containing  carbon  monoxide 
bright  red.  This  difference  disappears  slowly  but  not  entirely 
for  weeks, 

5.  Tannin  Test. — Mix  an  aqueous  blood  solution^  with  3  times 
its  volume  of  i  per  cent,  tannin  solution  and  shake  thor- 
oughly. A  difference  in  color  between  normal  and  carbon 
monoxide  blood  can  be  recognized  after  several  hours,  most 
distinctly  after  24  hours.  Normal  blood  is  gray  but  carbon 
monoxide  blood  is  crimson  red.  This  difference  is  apparent 
even  after  several  months.  Ten  per  cent,  of  carboxyhaemo- 
globin  can  be  detected  in  blood  by  tests  4  and  5. 

6.  Copper  Sulphate  Test. — A  drop  of  saturated  copper  sul- 
phate solution  added  to  2  cc.  of  carbon  monoxide  blood  mixed 
with  the  same  volume  of  water  gives  a  brick-red  precipitate. 
The  deposit  from  normal  blood  is  greenish  brown.  In  all  these 
precipitation  tests  (4,  5  and  6)  the  less  easily  decomposed 
carbon  monoxide  blood  remains  bright  red  but  the  more  easily 
decomposed  normal  blood  in  presence  of  the  precipitants  used 
and  others  is  off  color  or  dark. 

7.  Ammonium  Sulphide  Test. — Mix  0.2  cc.  of  ammonium 
sulphide  solution  and  0.2-0.3  cc.  of  30  per  cent,  acetic  acid  with 
ID  cc.  of  2  per  cent,  aqueous  blood  solution.  Carbon  monoxide 
blood  gives  a  fine  rose  color  but  normal  blood  is  greenish  gray. 
The  former  within  24  hours  gives  a  red  flocculent  precipitate. 

8.  Palladous  Chloride  Test. — Carbon  monoxide  precipitates 
black  metallic  palladium  from  a  neutral  aqueous  palladous 
chloride  solution: 

CO  +  H2O  +  PdCla  =  CO2  +  2HCI  +  Pd. 

^  Mix  I  volume  of  glacial  acetic  acid  with  2  volumes  of  water.  This  acid  con- 
tains about  30  per  cent,  acetic  acid. 

2  Use  I  part  of  blood  to  4  parts  of  water. 


DETECTION    OF    CAHIJON    MONOXIDE  BLOOD 


299 


Mix  a  few  drops  of  potassium  hydroxide  solution  with  the  blood 
and  warm  gently  upon  the  water-bath.  By  means  of  a  suction 
pump  draw  through  the  solution  air  that  has  been  washed  until 
pure.  Pass  the  gas  evolved  first  through  lead  acetate  solution 
to  remove  possible  hydrogen  sulphide,  then  through  sulphuric 
acid  to  absorb  ammonia  and  finally  through  a  neutral  light  red 
palladous  chloride  solution  (i  :5oo). 

9.  Spectroscopic  Examination. — The  detection  of  carboxy- 
haemoglobin  with  the  spectroscope  is  comparatively  easy.     The 


Oxy  haemoglobin. 


Haemoglobin. 


Methaemoglobin. 


Carboxyhaemoglobin. 


Haematoporphyrin,  very 
dilute,  acid. 


Haematoporphyrin,  not  so 
dilute,  alkaline. 


Fig.  24. — Absorption-Spectra. 


two  absorption  bands  of  this  compound  are  quite  similar  to 
those  of  oxyhasmoglobin  but  they  he  somewhat  nearer  together 
and  more  toward  the  violet.  The  main  difference,  however, 
between  the  absorption  bands  of  these  compounds  is  that  those 
of  carboxyh^moglobin  are  not  extinguished  by  reducing  agents. 
To  prepare  the  blood  solution  for  spectroscopic  examination, 
dilute  1-1.5  parts  of  blood  with  100  parts  of  water  and  make  the 


300  DETECTION   OE   POISONS 

observations  through  a  layer  i  cm.  thick.  To  reduce  i  per 
cent,  blood  solution,  mix  thoroughly  with  a  few  drops  of  am- 
monium sulphide  solution  and  add  4-6  drops  more  of  the  same 
reagent  as  a  surface  layer  to  exclude  air.  Reduction  begins  in 
about  6-8  minutes.  A  solution  of  tartaric  acid  and  ferrous 
sulphate  in  presence  of  an  excess  of  ammonium  hydroxide 
solution  will  also  reduce  oxyhaemoglobin. 

Oxyhaemoglobin  under  these  conditions  is  changed  to  reduced 
haemoglobin.  The  two  absorption  bands  characteristic  of  the 
former  disappear  and  a  broad  diffuse  absorption  band  occupies 
the  previous  bright  space  between  the  two  bands.  The  spectrum 
of  carboxyhsemoglobin  remains  unchanged  only  when  27  per 
cent,  at  least  of  the  haemoglobin  is  saturated  with  carbon 
monoxide.  If  allowed  to  stand  in  an  open  vessel,  blood  will  lose 
carbon  monoxide  within  8  days.  But  carbon  monoxide  blood 
sealed  in  glass  tubes  is  said  to  keep  for  years.  Carbon 
monoxide  has  been  detected  in  blood  of  a  cadaver  after  18 
months. 

ToUens^  recommends  adding  some  formaldehyde  to  the  blood  solution.  This 
reagent  has  not  the  slightest  effect  upon  the  two  oxyhemoglobin  bands.  Warm- 
ing the  mixture  very  gently  with  ammonium  sulphide  solution  develops  a  third 
and  nearly  as  distinct  black  band  almost  midway  between  the  original  bands 
which  gradually  disappear.  Finally  only  this  band  wiU  remain.  This  is  a  far 
more  satisfactory  test  than  that  given  by  the  indefinite  band  of  blood  alone. 
If  the  solution  is  cooled  and  agitated  with  air,  this  third  band  will  disappear  and 
the  two  original  oxyhaemoglobin  bands  will  return. 

If  carbon  monoxide  is  present,  formaldehyde  does  not  have  this  action. 

2.  Detection  of  Blood  Stains 

The  detection  of  blood  in  dry  stains  upon  fabrics,  wood,  knives, 
weapons,  etc. ,2  is  more  certain  and  less  open  to  question,  if 
haemiin  crystals  (Teichmann's  blood  crystals)  are  prepared  from 
the  blood  pigment.  If  haemin  crystals  are  obtained,  the  stain 
in  question  may  be  regarded  with  certainty  as  due  to  blood. 
Fresh  blood  when  dry  is  bright  red  and  has  a  smooth  surface. 

^  Berichte  der  Deutschen  chemischen  Gesellschaft  34,  1426  (1901). 
2  Blood  mixed  with  iron  oxide  as,  for  example,  blood  upon  rusty  knives  and 
weapons  usually  fails  to  give  hasmin  crystals. 


DETECTION   OF   CARJ^ON   MONOXIDE  BLOOD  301 

Flakes  of  such  blood  scraped  from  any  material  arc  garnet  red 
by  transmitted  light.  A  solution  of  fresh  blood  stains  in  potas- 
sium or  sodium  hydroxide  is  dichroic,  being  red  by  transmitted 
and  green  by  reflected  light.  Later  dried  blood  becomes 
brownish  red  or  dark  brown.  These  color  changes  are  due  to 
conversion  of  oxyhaemoglobin  into  methsemoglobin  and  then 
into  hsematin.  The  first  two  substances  are  soluble  in  water 
but  the  last  is  not.  But  haematin  is  soluble  in  alkalies  and  in 
alcohol  containing  sulphuric  acid.  This  change  of  the  blood 
pigment  depends  not  only  upon  the  age  of  the  stain  but  really 
upon  the  action  of  air  (oxygen),  light,  heat  and  moisture  upon 
the  blood  before  it  is  dry.  If  the  blood  is  in  a  thin  layer, 
haemoglobin  will  sometimes  change  into  methaemoglobin  even 
in  3-10  days.  BoiUng  water  causes  immediate  insolubility. 
The  action  is  also  very  rapid  in  direct  sunhght.  Washing  in 
alkaline  solutions  (boiling  solutions  of  potassium  or  sodium 
soap,  sodium  carbonate  solution,  ammonia  and  sewage)  also 
causes  rapid  decomposition.  But  acids,  nitric  and  hydrochloric, 
as  well  as  putrefaction,  act  more  slowly,  giving  the  blood  a 
laked  appearance  and  even  making  it  clear  and  colorless.  If, 
however,  the  blood  has  once  dried,  these  injurious  agencies, 
even  putrefaction,  act  less  easily. 

Preparation  of  Haemin  Crystals. — Prepare  a  cold  aqueous 
extract  of  the  stain  as  free  as  possible  from  fibers  and  evaporate 
the  solution  upon  a  watch  glass  away  from  dust.  Add  a  trace 
of  sodium  chloride^  to  the  residue,  also  8-10  drops  of  glacial 
acetic  acid,  and  stir  with  a  glass  rod.  Heat  just  for  an  instant 
over  a  small  flame,  then  evaporate  the  solution  gradually 
upoh  a  moderately  warm  water-bath  and  examine  the  residue 
with    a   microscope    magnifying    300-500    times.     If    hasmin 

1  Strzyzowski  (Chemisches  Centralblatt,  1897,  I,  295)  advises  using  sodium 
iodide  instead  of  sodium  chloride.  Place  a  small  particle  of  material  suspected 
of  containing  blood  upon  a  glass  slide  and  add  a  drop  of  sodium  iodide  solution 
(i  :  500).  Evaporate  and  cover  with  a  cover-glass.  Heat  for  3-6  seconds 
with  concentrated  acetic  acid  which  is  allowed  to  run  under  the  cover-glass.  The 
test  with  this  modification  is  said  to  be  more  deHcate,  o\\-ing  to  the  darker  color 
of  the  hasmatin  hydriodide  crj'stals.  The  cr\'stals  are  usually  obtained  in  less 
time  and  with  as  small  a  quantity  as  0.000025  gram  of  fresh  blood.     Tr. 


302  DETECTION   OF   POISONS 

crystals  fail  to  appear,  repeat  the  evaporation  several  times, 
using  in  each  instance  8-10  drops  of  glacial  acetic  acid,  and 
examine  the  residue  each  time  under  the  microscope.  Hsemin 
crystals  are  brownish  red  to  dark  brown  and  form  rhombic  scales 
which  frequently  lie  crossed  (Fig.  25).     Usually  glacial  acetic 

acid  is  the  only  solvent  that  will  ex- 
tract the  pigment  from  old  blood 
stains.  Brucke  heats  the  stains  or 
scrapings  to  boiling  in  a  test-tube 
with  10-20  drops  of  glacial  acetic 
acid.  The  decanted  or  filtered  solu- 
tion, after  addition  of  a  trace  of 
sodium  chloride,  is  evaporated  upon 
a  watch  glass  to  dryness  at  40-80° 
Fig.  25.— Hffimin  Crystals.       ^^^   the  residue  is  examined  under 

the  microscope.     By  this  method  it 
is  immaterial  whether  the  blood  has  coagulated  or  not. 

Cold  water  is  without  effect  upon  blood  stains,  if  they  have 
previously  been  treated  with  hot  water.  Protein  substances  in 
the  blood  are  thus  coagulated  and  rendered  insoluble.  In 
such  a  case  treat  the  stain  with  water  containing  a  few  drops  of 
sodium  hydroxide  solution.  If  the  stains  are  upon  wool,  use 
very  dilute  sodium  hydroxide  solution  since  alkalies  dissolve 
wool.  Water  containing  ammonium  hydroxide  will  extract 
stains  and  this  alkali  does  not  act  upon  wool.  Use  the  alkaline 
aqueous  extract  to  prepare  hasmin  crystals.  Evaporate  the 
solution  to  dryness  in  a  watch  glass  upon  the  water-bath  and 
mix  the  residue  intimately  with  8-10  drops  of  glacial  acetic  acid. 
Add  a  trace  of  sodium  chloride  and  again  evaporate.  Sometimes 
it  is  advisable,  after  acidifying  the  extract  of  the  stain  with 
acetic  acid,  to  add  tannic  acid,  or  zinc  acetate,  and  prepare 
Teichmann's  crystals  from  the  precipitate. 

Occasionally  it  is  necessary  to  extract  suspected  stains  with 
hot  alcohol  containing  sulphuric  acid.  Hsematin  formed  from 
the  blood  pigment  dissolves.  If  this  compound  is  present,  the 
solution  has  a  brown  color.  Excess  of  sodium  hydroxide  solu- 
tion will  produce  the  dichroism  characteristic  of  an  alkaline 


DETECTION    OF   CARBON    MONOXIDE   BLOOD  303 

haematin  solution,  namely,  red  by  transmittec]  and  green  by 
reflected  light.  Obviously,  ha}matin  should  be  identified  by 
the  spectroscope  both  in  acid  and  alkaline  solution. 

Blood  mixed  with  iron  oxide  (blood  upon  rusty  weapons) 
usually  fails  to  give  haemin  crystals  but  the  extract  with  dilute 
sodium  hydroxide  solution  frequently  shows  the  dichroism  of 
haematin  solution.  Since  iron  oxide  or  rust  forms  an  insoluble 
compound  with  haematin,  warm  such  stains  for  some  time  upon 
the  water-bath  with  sodium  hydroxide  solution  to  dissolve  any 
haematin  present. 

Haematin. — Warming  an  aqueous  blood  solution  to  about  70°  decomposes  the 
blood  pigment  oxyhccmoglobin  into  a  protein  substance  called  globin  and  haematin 
a  pigment  containing  iron.  Acids,  alkalies  and  several  metallic  salts  decompose 
oxyhaemoglobin  in  the  same  way.  If  this  decomposition  takes  place  in  the  ab- 
sence of  oxygen,  another  pigment  appears.  Hoppe-Seyler  gave  the  latter  the 
name  hsemochromogen  and  other  experimenters  have  called  it  "reduced  haematin." 
Oxygen  and  consequently  air  rapidly  oxidizes  this  pigment  to  haematin.  On  the 
other  hand  reducing  agents  like  ammonium  sulphide  convert  haematin  into  hasmo- 
chromogen.  Different  formulas  are  given  for  haematin.  W.  Kiister  and  others 
now  give  it  the  formula  C34H34N4Fe06.  Haematin  is  amorphous  and  has  a  dark 
brown  or  blue-black  color.  In  water,  dilute  acids,  alcohol,  ether  and  chloroform 
it  is  insoluble  but  soluble  in  alcohol  or  ether  containing  acid.  In  even  very  dilute 
solutions  of  caustic  alkalies  it  is  freely  soluble.  Alkaline  haematin  solutions  are 
dichroic.  In  rather  thick  layers  the  color  appears  red  by  transmitted  light  and 
greenish  in  thin  layers.  Acid  solutions  are  always  brown.  Alkaline  haematin 
solutions  are  precipitated  by  calcium  or  barium  hj^droxide  solution. 

Haemin  is  the  hydrochloric  ester  of  haematin.  Very  prob- 
ably haemin  has  the  empirical  formula  C34H33N4Fe04Cl. 

Note. — If  the  blood  stain  is  perfectly  fresh,  it  may  be  recog- 
nized by  observing  blood-corpuscles  with  the  microscope. 
Human  blood  can  be  differentiated  from  animal  blood  by  com- 
paring blood-corpuscles  with  those  of  animal  blood  as  to  size, 
only  when  the  corpuscles  are  still  intact. 

Spectroscopic  Detection  of  Blood 

If  the  extract  of  a  blood  stain  with  cold  water  is  already 
brown,  a  third  fainter  and  narrower  band  will  appear  in  ad- 
dition to  the  two  oxyhaemoglobin  bands.  This  Hes  in  the 
orange  between  C  and  D   and  is   the  methaemoglobin  band 


304  DETECTION   OF  POISONS 

Cold  water  will  dissolve  most  of  the  methgemoglobin  from 
fresh,  dried  blood  stains. 

Acetic  acid  will  discharge  these  two  bands,  if  the  oxyhaemo- 
globin  solution  is  not  too  dilute.  At  the  same  time  the  solution 
will  become  mahogany-brown  from  formation  of  haematin  in 
acid  solution.  This  solution  has  a  characteristic  gpectrum, 
namely,  four  absorption  bands  in  the  yellow  and  green.  If 
excess  of  ammonium  hydroxide  is  added  to  this  solution,  the 
alkaline  solution  contains  haematin,  recognizable  by  a  broad 
faint  absorption  band  lying  between  the  red  and  yellow.  A 
few  drops  of  ammonium  sulphide  solution  will  extinguish  this 
band  and  bring  out  two  broad  bands,  namely,  one  in  the  green 
and  the  other  in  the  light  blue.  These  bands  lie  farther  to  the 
right  than  do  those  of  oxyhasmoglobin  and  are  of  about  the 
same  width.  This  is  the  spectrum  of  reduced  haematin  (haemo- 
chromogen).  All  these  spectroscopic  tests  are  very  charac- 
teristic, especially  the  spectra  of  oxyhaemoglobin,  haemoglobin 
and,  in  the  case  of  old  blood  stains,  that  of  reduced  haematin. 

Up  to  the  present  time  no  red  solution  has  been  found  which, 
upon  abstraction  and  addition  of  oxygen,  will  give  the  same 
spectroscopic  phenomena  as  blood. 

When  the  quantity  of  blood  is  very  small,  or  when  the  blood 
pigment  has  undergone  further  decomposition,  so  that  the 
bands  of  oxyhaemoglobin  are  no  longer  visible,  it  is  advisable 
to  extract  the  stains  for  several  hours  with  concentrated  potas- 
sium cyanide  solution.  Blood  will  give  a  light  red  or  yellowish 
brown  solution  containing  the  cyano-compound  of  haematin. 
The  spectrum  of  hcematin  in  alkaline  solution  will  appear  as  a 
broad,  faint  band. 

The  investigations  of  Kratter  and  Hammerl  have  shown  that 
charred  blood,  which  no  longer  responds  to  any  of  the  other 
blood  reactions,  will  still  give  the  haematoporphyrin  spectrum 
upon  treatment  with  concentrated  sulphuric  acid  (E.  v.  Hof- 
mann,  Lehrbuch  der  gerichtlichen  Medizin,  1903). 

Ammoniacal  carmine  solution  gives  two  absorption  bands 
similar  to  those  of  oxyhaemoglobin  but  they  do  not  change 
upon  addition  of  acetic  acid  or  ammonium  sulphide.     A  band 


DETECTION   OF   CARBON    MONOXIDE   BLOOD  305 

given  by  fuchsine  analogous  to  that  of  haemoglobin  remains 
unchanged  after  agitation  with  air. 

Other  Blood  Tests 

I.  Schonbein-Van  Deen  Ozone  Test. — A  mixture  of  ozon- 
ized turpentine^  and  alcohoHc  tincture  of  guaiac  resin,  shaken 
with  a  little  blood,  produce  a  hght  blue  color.  Separated  from 
the  turpentine,  the  tincture  is  deep  blue.  Though  very 
dehcate,  this  test  is  not  characteristic  of  blood,  for  many 
iriorganic  and  organic  substances  under  the  same  conditions 
produce  "guaiac  blue."  Nitrous  acid,  chlorine,  bromine  and 
iodine,  chromic  and  permanganic  acids,  ferric  and  cupric  salts 
produce  blue  solutions  direct  with  guaiac  resin.  In  examining 
blood  stains  usually  it  is  possible  to  exclude  these  substances 
beforehand.  But  other  substances  hke  cell  contents  or  haemo- 
globin, having  the  power  of  transferring  ozone,  may  attach 
false  significance  to  the  guaiac-blue  reaction.  Enzjones 
(diastases),  hydrolytic  ferments  (enzymes  in  the  narrower 
sense),  as  well  as  the  so-called  oxidation  ferments  (oxidases),  are 
organic  substances  of  this  character.  They  occur  in  different 
parts  of  plants,  especially  in  fungi  and  in  seeds.  Saliva,  ex- 
tracts of  certain  organs,  contents  of  white  blood-corpuscles  and 
pus  cells  are  animal  products  of  similar  nature.  E.  Schaer- 
states  that  these  animal  and  vegetable  substances  differ  from 
hydrogen  dioxide  in  being  catalytic  in  action  and  carriers  of  oxy- 
gen at  the  same  time.  And  also  that  a  temperature  of  ioo°, 
or  contact  with  hydrocyanic  acid,  completely  destroys  their 
power  of  transferring  oxygen,  or  at  least  greatly  diminishes  it.  in 
which  respect  they  are  essentially  different  from  haemoglobin. 
Neither  high  temperature  (ioo°)  nor  hydrocyanic  acid  has  any 
restraining  influence  upon  haemoglobin  so  far  as  transference  of 
oxygen  is  concerned.  Consequently  an  extract,  containing  one 
of  these  ferment-like  substances  but  no  blood,  placed  even  for  a 

^  Turpentine  always  contains  ozone,  if  exposed  to  light  for  a  long  time  in  a 
loosely  stoppered  bottle. 

2  Forschungsberichte  iiber  Nahrungsmittel,  etc.,  3,  i  (1S96)  and  Archiv  der 
Pharmazie  236,  571  {li 
20 


306  DETECTION   OP   POISONS 

short,  time  in  a  hot  water-bath,  loses  the  power  of  giving  the 
"guaiac  blue"  test.  In  absence  of  blood,  the  result  will  also 
be  negative,  if  the  extract  of  the  suspected  stain  is  treated  with 
hydrocyanic  acid.  For  these  reasons  great  care  is  necessary  in 
interpreting  a  positive  guaiac  test  given  by  the  extract  of  a 
supposed  stain.  The  gaaiac  test  is  certainly  very  useful  as  a 
delicate  preliminary  test  and  in  many  instances  as  a  check  upon 
blood.  The  three  forms  of  the  blood  pigment  entering  into 
such  an  examination,  namely,  hgemoglobin,  methaemoglobin 
and  haematin,  are  alike  in  the  guaiac  test,  at  least  quahtatively, 
as  far  as  transference  of  oxygen  is  concerned.  The  examination 
and  extraction  of  the  stain  may,  therefore,  be  conducted  in 
neutral,  acid  or  alkaline  solution,  depending  upon  the  nature  of 
the  substance,  and  either  hot  or  cold.  Render  an  alkaline 
extract  faintly  acid  with  acetic  acid  before  adding  guaiac  tinc- 
ture. In  many  instances  it  is  advisable  to  extract  the  blood 
stain  with  hot  alcohol  containing  sulphuric  acid.  Treat  such  an 
acid,  alcoholic  haematin  solution  with  guaiac  tincture  direct. 
Addition  of  water  will  precipitate  the  resin  with  the  adherent 
blood  pigment. 

(a)  Vitali's  Procedure. — Extract  the  stain  with  water  con- 
taining carbon  dioxide,  or  old  stains  with  very  dilute  sodium 
hydroxide^  solution  free  from  nitrite  and  nitrate.  Filter  the 
extract  and  add  a  little  alcoholic  guaiac  tincture  to  a  portion  of 
the  filtrate  after  acidification  with  acetic  acid,  if  necessary. 
If  the  milky  liquid  is  not  blue  in  15  minutes,  interfering  oxidizing 
agents  are  absent.  Then  add  a  few  drops  of  old  turpentine  and 
shake.  The  milky  liquid  will  turn  blue  at  once,  or  in  a  short 
time,  if  blood  pigment  is  present.  Very  gentle  warming  upon 
the  water-bath  increases  the  delicacy  of  the  reaction.  Even 
putrid  blood  2  months  old  is  said  to  give  a  positive  test. 

(b)  E.  Schaer's  Procedure.— Blood  stains  upon  linen,  though 
quite  old,  dissolve  completely  when  treated  for  some  time  with 
70  per  cent,  chloral  hydrate  solution.  Moistening  the  stains 
beforehand  with  glacial  acetic  acid  aids  solution.  Also  pre- 
pare an  extract  of  guaiac  resin  in  70  per  cent,  chloral  hydrate 

^  Use  sodium  hydroxide  prepared  from  metallic  sodium  in  this  test. 


DETECTION   OF   CARBON    MONOXIDE   BLOOD  .'i07 

solution.  Mix  the  extract  of  the  stain  with  an  equal  volume  of 
the  latter  solution.  In  absence  of  nitrites,  the  color  of  this 
mixture  is  brownish  yellow  to  light  brown.  If  preferred,  a 
contact  test  for  blood  may  be  made  by  this  method.  Add  to 
the  mixture  of  blood  and  guaiac  Hiinefeld's^  turpentine  solu- 
tion, or  hydrogen  peroxide,  as  a  surface-layer.  An  intense 
blue  zone  will  appear  where  the  two  solutions  meet.  Guaiaconic 
acid  in  guaiac  resin  produces  "guaiac  blue."  O.  Dobner  has 
suggested  substituting  a  dilute  solution  of  guaiaconic  acid  for 
guaiac  resin.  Blood,  or  blood  pigment,  behaves  like  a  ferment 
and  activates  the  ozonized  turpentine  or  hydrogen  peroxide, 
either  of  which  by  itself  will  not  turn  the  solution  of  guaiac 
resin  blue. 

2.  Schaer's  Aloin  Test. — The  same  conditions,  producing 
"guaiac  blue"  from  guaiaconic  acid,  give  rise  to  "aloin  red" 
from  aloin.  This  substance  has  a  stronger  coloring  power  and 
lasts  longer  than  "guaiac  blue."  Use  the  same  solution  of  blood 
in  70-75  per  cent,  chloral  hydrate  solution  mixed  with  a  weak 
chloral  hydrate  solution  of  aloin.  Add  Hiinefeld's  hydrogen 
peroxide  solution  as  a  surface  layer.  After  some  time  a  violet- 
red  zone  will  appear  and  a  red  color  of  equal  intensity  will 
gradually  extend  throughout  the  aloin  solution.  Another 
method  of  making  this  test  consists  in  first  extracting  the  blood 
stain  with  pure  water,  acetic  acid,  chloral  hydrate  solution  or 
alkaline  salt  solution.  Neutralize  this  solution  and  add  dilute 
alcoholic  aloin  solution  and  hydrogen  peroxide.  If  the  sus- 
pected stain  contains  blood  pigment,  a  red  color  will  appear  at 
once  and  persist  for  a  long  time. 

3.  Biological  Detection  of  Hiiman  Blood^ 

Injection  of  bacteria  produces  specific,  bacteriolytic  bodies 
and  similarly  injection  of  the  blood  of  one  animal  species  into 

^  See  page  314  for  the  preparation  of  tliis  reagent. 

2  This  subject  has  been  introduced  for  the  sake  of  completeness.  If  such  an 
investigation  is  for  forensic  purposes,  the  chemist  will  either  decline  to  undertake 
it,  or  conduct  the  experiment  with  an  associate  who  has  had  bacteriological  and 
pathological  experience. 


308  DETECTION   OP   POISONS 

an  animal  of  a  different  species  gives  rise  to  specific,  haemolytic 
and  agglutinating  bodies.  Rabbit's  blood,  for  example,  in- 
jected repeatedly  into  a  guinea  pig,  develops  in  the  serum  of 
such  a  guinea  pig  substances  capable  of  agglutinating  and  dis- 
solving red  corpuscles  of  the  rabbit,  setting  haemoglobin  free 
and  rendering  the  blood  laky.  Blood  serum  from  an  animal, 
into  which  defibrinated  blood,  or  blood  serum  from  a  different 
animal  species  has  been  injected  intravenously,  subcutaneously 
or  intraperitoneally,  that  is  to  say,  into  the  peritoneal  cavity, 
has  the  peculiar  property  of  causing  precipitation  only  in  blood 
serum  of  this  particular  animal  species.  Uhlenhuth,^  Wasser- 
mann  and  Schiitze,^  and  others  have  made  independent  experi- 
ments of  this  kind  with  blood  serum  to  find  for  forensic  purposes 
a  test,  based  upon  this  biological  method,  which  shall  differenti- 
ate human  blood  from  the  blood  of  every  other  animal  species. 
Repeated  injection  of  lo  cc.  of  defibrinated  human  blood,  or 
human  blood  serum  free  from  cells,  into  a  rabbit,  either  intra- 
peritoneally or  subcutaneously,  yields  a  serum  producing  a 
heavy,  cloudy  precipitate  in  an  aqueous  solution  of  human  blood. 
This  coagulin  is  specific  in  action,  producing  a  precipitate  only 
in  presence  of  human  blood.  Wassermann  and  Schiitze  tested 
the  blood  of  23  different  animals,  among  which  were  mammals, 
birds  and  fishes,  and  obtained  negative  results  with  blood 
solutions  from  these  very  different  animal  species.  By  use  of 
blood  serum  it  is  possible  to  differentiate  even  old  human  blood, 
dried  for  many  weeks,  from  the  blood  of  other  animals. 

To  demonstrate  the  use  of  this  method,  A.  Dieudonne^ 
prepares  i  per  cent,  blood  solutions,  placing  2  cc.  of  the  clear 
filtered  solution  in  small  test-tubes  and  adding  an  equal  volume 
of  double  physiological  salt  solution  (=1.8  per  cent.  NaCl). 
Then  add  6  drops  of  serum  to  each  portion  and  place  the  tubes 
in  an  incubator  at  37°.  The  serum  of  the  rabbit,  treated  with 
human  blood  serum,  added  to  an  aqueous  solution  of  human 

^Deutsche  medizinische  Wochenschrift,  1901,  No.  6;  und  Zeitschrift  fiir 
Medizinalbeamte,  1903,  Heft  5  and  6. 

^Berliner  klinische  Wochenschrift,  1901,  No.  7. 
^Munchener  medizinische  Wochenschrift,  1901,  page  533. 


DETECTION  OF  CARBON  MONOXIDE  BLOOD       309 

blood,  produced  in  a  few  minutes  a  distinct  flocculent  precipi- 
tate which  gradually  became  more  and  more  marked.  As  a 
check,  test  also  with  normal  rabbit's  serum  which  will  cause  no 
precipitate  in  a  solution  of  human  blood.  Dieudonne  found  also 
that  rabbit's  serum,  obtained  after  injecting  human  blood  serum, 
causes  precipitates  not  only  in  human  blood  solutions  but  in 
human  urine  containing  albumin,  with  an  exudate  from  human 
pleura  and  with  peritoneal  exudate.  But  precipitation  in  the 
case  of  human  blood  was  much  more  marked  than  in  these  other 
tests.  In  his  experiments  Dieudonne  used  blood  expressed 
from  the  placenta,  repeatedly  injecting  it  subcutaneously  into 
rabbits  in  separate  doses  of  lo  cc.  and  at  intervals  of  3-4  days. 
The  animals  were  bled  several  days  after  the  last  injection  and 
the  blood  was  kept  upon  ice. 

The  antiserum  used  in  detecting  blood  should  above  every- 
thing else  be  perfectly  clear.  To  prepare  such  serum,  use  a 
sterile  Berkefeld  filter  attached  to  a  water  pump.  The  anti- 
serum should  be  active  in  very  dilute  solution.  Distinct  tur- 
bidity should  appear  immediately  in  a  solution  diluted  i  :  1000, 
or  in  1-2  minutes  at  latest.  Sera  must  be  of  this  high  efficiency 
for  practical  use.  Uhlenhuth  has  shown  that  the  biological 
method  of  detecting  blood  is  specific  for  human  albumin.  A 
necessary  consequence  of  this  fact  is  that  the  material  should 
first  of  all  be  shown  to  be  blood.  The  first  question  for  the 
expert  to  answer  in  such  an  investigation  must  always  be: 
''Is  there  any  blood  at  all  present?"  If  the  answer  is  afiirma- 
tive,  the  next  question  is:  "Is  it  human  or  animal  blood?" 
Consequently  the  material  should  first  be  examined  for  blood 
stains  by  van  Deen's  ozone  test,  Teichmann's  haemin  test  and 
by  the  spectroscope.  If  the  suspected  stains  are  upon  a  hard 
surface,  as  a  knife,  hatchet,  gun  barrel,  wood,  stone,  etc.,  they 
should  be  scraped  off  for  the  biological  blood  test  and  extracted 
for  several  hours  in  a  test-tube  with  physiological  salt  solution 
(=  0.9  per  cent.  NaCl).  First,  filter  the  extract  through  paper. 
If  the  filtrate  is  not  clear,  next  use  a  Berkefeld  filter. 


APPENDIX 
PREPARATION  OF  REAGENTS 

General  Alkaloidal  Reagents. — ^A  class  of  reagents,  known  as 
general  alkaloidal  reagents,  added  to  solutions  of  most  of  the 
alkaloids  or  of  their  salts,  produce  precipitates  characterized  by 
their  color,  their  amorphous  or  crystalline  appearance  and  their 
insolubility  or  sparing  solubility  in  water.  But  these  reagents 
do  not  precipitate  alkaloids  exclusively.  Several  members  of 
this  class,  for  example,  the  chlorides  of  gold,  platinum  and 
mercury,  phospho-molybdic  and  phospho-tungstic  acids,  react 
similarly  with  ammonia  and  many  ammonium  derivatives.  An 
explanation  of  this  similarity  in  behavior  is  found  in  the  fact 
that  most  of  the  alkaloids,  being  secondary  or  tertiary  bases,  are 
themselves  ammonium  derivatives.  Nearly  all  the  general 
alkaloidal  reagents  also  precipitate  proteins,  albumoses,  pep- 
tones, creatinine  and  the  nuclein  bases,  adenine,  guanine,  hy- 
poxanthine  and  xanthine. 

The  general  alkaloidal  reagents  are  especially  useful  in 
detecting  the  presence,  or  absence,  of  alkaloids  and  other  basic 
compounds.  If  there  is  only  a  slight  residue  from  the  ether 
extract  of  the  alkaline  solution  in  the  Stas-Otto  method,  test 
first  with  the  general  alkaloidal  reagents  and  then,  if  neces- 
sary, for  individual  alkaloids.  To  perform  these  tests,  dissolve 
the  given  residue  in  very  dilute  hydrochloric  or  sulphuric  acid, 
distribute  the  filtered  solution  upon  several  watch  glasses  and 
add  to  each  portion  a  drop  of  the  more  sensitive  reagents. 
If  an  alkaloid  or  any  other  basic  substance  is  present,  distinct 
precipitates  or  at  least  decided  cloudiness  will  appear  in  all  or 
in  nearly  all  of  the  tests. 

The  most  important  general  alkaloidal  reagents  are  the 
following : 

Gold  Chloride  dissolved  in  water  (i  -.30)  produces  white,  yel- 
low  or  brown  precipitates  which  are  amorphous  or  crystal- 

310 


PREPAKATION    OF    REAGENTS  .'ill 

line.  These  precipitates  decompose  to  some  extent  with 
separation  of  metallic  gold. 

Platinum  Chloride  dissolved  in  water  (i  :  20)  produces  yellow- 
ish white  to  yellow  precipitates  which  are  usually  granular  and 
crystalline.  These  precipitates  are  usually  analogous  in  com- 
position to  ammonium  chloroplatinate,  (H4N)2PtCl6. 

Mercuric  Chloride  dissolved  in  water  (i  :  20)  produces  white 
to  yellowish  precipitates  which  are  usually  amorphous  but 
gradually  become  crystalline. 

lodo-potassium.  Iodide,  prepared  by  dissolving  5  parts  of 
iodine  and  10  parts  of  potassium  iodide  in  100  parts  of  water, 
produces  brown  precipitates  which  are  usually  flocculent. 

Potassium  Cadmitmi  Iodide,  prepared  by  dissolving  20  grams 
of  potassium  iodide  in  20  cc.  of  boiling  water,  adding  10  grams 
of  cadmium  iodide  and  diluting  to  100  cc,  produces  white  or 
yellowish  precipitates  with  sulphuric  acid  solutions  of  most  of 
the  alkaloids,  even  when  these  solutions  are  very  dilute.  These 
precipitates,  at  first  amorphous  but  later  crystalline,  dissolve 
in  an  excess  of  the  reagent  and  also  in  alcohol. 

Potassitun  Bismuthous  Iodide  may  be  prepared  according  to 
Kraut^  by  dissolving  80  grams  of  bismuth  subnitrate  in  200 
grams  of  nitric  acid  (sp.  gr.  1.18  =  30  per  cent.  HNO3)  and  pour- 
ing this  solution  into  a  concentrated  solution  of  272  grams  of 
potassium  iodide  in  water.  Allow  the  potassium  nitrate  to 
crystallize  and  dilute  the  solution  with  water  to  1000  cc.  This 
reagent  produces  orange-red  precipitates  with  sulphuric  acid 
solutions  of  many  alkaloids.  By  shaking  these  precipitates  with 
sodium  hydroxide  and  carbonate  solution,  it  is  often  possible 
to  recover  the  alkaloids  unchanged  and  sometimes  almost 
quantitatively. 

Potassium  Mercuric  Iodide,  prepared  by  dissolving  1.35 
grams  of  mercuric  chloride  and  5  grams  of  potassium  iodide  in 
100  cc.  of  water,  produces  white  or  yellowish  precipitates  with 
hydrochloric  acid  solutions  of  most  of  the  alkaloids.  These 
precipitates  at  first  amorphous,  gradually  become  crystalliiie. 

^  Annalen  der  Chemie  und  Pharmazie,  :?io,  310  (1SS2)  und  Archiv  der  Phar- 
mazie,   235,    152    (1897). 


312  DETECTION   OF   POISONS 

Potassium  Zinc  Iodide  is  prepared  by  dissolving  lo  grams  of 
zinc  iodide  and  20  grams  of  potassium  iodide  in  100  cc.  of  water. 

Phospho-molybdic  Acid  may  be  prepared  by  either  of  the 
following  methods : 

(a)  Saturate  sodium  carbonate  solution  with  pure  molybdic 
acid,  add  i  part  of  crystallized  disodium  phosphate  (Na2HP04.- 
12H2O)  to  5  parts  of  the  acid  and  evaporate  to  dryness.  Fuse 
the  residue  in  a  porcelain  crucible  and  dissolve  the  cold  melt  in 
water.  Prepare  10  parts  of  solution  from  i  part  of  this  residue. 
Add  enough  nitric  acid  to  the  filtered  solution  to  produce  a 
golden  yellow  color. 

(b)  If  molybdic  acid  is  not  at  hand,  completely  precipitate  at 
40°  with  excess  of  sodium  phosphate  solution  the  nitric  acid 
solution  of  ammonium  molybdate  used  in  testing  for  phos- 
phoric acid.  Thoroughly  wash  the  yellow  precipitate,  add 
water  and  dissolve  in  warm  concentrated  sodium  carbonate 
solution.  Evaporate  this  solution  to  dryness  and  fuse  the  resi- 
due until  ammonia  is  completely  expelled.  If  there  is  any 
reduction  (blue  or  black  color),  moisten  the  residue  with  nitric 
acid  and  fuse  again.  Dissolve  this  residue  in  hot  water  and 
add  nitric  acid  in  large  excess.  Prepare  10  parts  of  solution 
from  I  part  of  residue.  The  golden  yellow  solution  should  be 
protected  from  ammonia  vapor. 

Phospho-molybdic  acid  produces  yellowish,  amorphous  pre- 
cipitates with  sulphuric  acid  solutions  of  most  of  the  alkaloids. 
After  a  while  these  precipitates  are  frequently  greenish  or 
bluish  from  reduction  of  molybdic  acid  to  molybdic  oxide. 

Phospho-tungstic  Acid,  prepared  by  adding  a  little  20  per 
cent,  phosphoric  acid  to  an  aqueous  solution  of  sodium  tung- 
state,  produces  precipitates  similar  to  those  given  by  phospho- 
molybdic  acid. 

Tannic  Acid  is  a  5  per  cent,  aqueous  solution  of  tannin.  This 
reagent  produces  whitish  or  yellowish,  flocculent  precipitates 
partially  soluble  in  hydrochloric  acid.  Alkaloids  may  be 
recovered  in  part  from  these  precipitates  by  treating  them  with 
lead  or  zinc  carbonate,  evaporating  to  dryness  and  extracting 
the  residue  with  ether,  alcohol  or  chloroform. 


PREPARATION  OF  REAGENTS  313 

Picric  Acid  is  a  concentrated  aqueous  solution  of  picric  acid 
which  produces  yellow  crystalline  precipitates,  or  amorphous 
precipitates  which  soon  become  crystalline. 

Picrolonic  Acid  is'  used  as  o.i  normal  alcoholic  solution  by 
dissolving  26.4  grams  of  solid  picrolonic  acid  (C10H8N4O5)  in  a 
liter  of  alcohol.  With  most  of  the  alkaloids  this  solution  pro- 
duces salts  called  picrolonates  which  are  crystalline,  difficultly 
soluble  and  yellow  to  red  in  color.  Picrolonic  acid  behaves  to- 
ward bases  like  a  monobasic  acid.^ 

B.  Other  Reagents  and  Solutions 

Erdmaiin's  Reagent. — Sulphuric  acid  containing  nitric  acid, 
prepared  by  adding  to  20  cc.  of  pure  concentrated  sulphuric 
acid  10  drops  of  a  solution  of  6  drops  of  concentrated  nitric  acid 
in  100  cc.  of  water. 

Froehde's  Reagent. — A  solution  of  molybdic  acid  in  sulphuric 
acid,  prepared  by  dissolving  5  mg.  of  molybdic  acid,  or  sodium 
molybdate,  in  i  cc.  of  hot,  pure  concentrated  sulphuric  acid. 
This  solution,  which  should  be  colorless,  does  not  keep  long. 

Fehling's  Solution. — The  two  following  solutions,  which 
should  be  kept  separate,  are  used  in  preparing  this  reagent: 

1.  Copper  Sulphate  Solution. — Dissolve  34.64  grams  of  pure 
crystallized  copper  sulphate  (CUSO4.5H2O)  in  sufficient  water  to 
make  500  cc. 

2.  Alkaline  Rochelle  Salt  Solution. — Dissolve  173  grams  of 
Rochelle  salt  (K.Na.C4H406.4H20)  and  50  grams  of  sodium 
hydroxide  in  hot  water  and  dilute  this  solution  when  cold  to 
500  cc. 

These  two  solutions,  mixed  volume  for  volume,  constitute 
Fehling's  solution  which  should  be  prepared  just  before  being 
used.  Fehling's  solution,  which  has  been  made  up  and  kept, 
should  always  be  tested  before  being  used.  The  solution  should 
not  be  used,  if  it  gives  a  red  precipitate  of  cuprous  oxide  when 
warmed  by  itself. 

^L.  Knorr,  Berichte  der  Deutschen  chemischen  Gesellschaft,  30,  914  (1897); 
H.  !Matthes  and  0.  Rammstedt,  Zeitschrift  fiir  analytische  Chemie  46,  565  and 
Archiv  der  Pharmazie  245,  112  (1907). 


314  DETECTION   OF   POISONS 

Formaldehyde -sulphuric  Acid. — Add  2-3  drops  of  aqueous 
formaldehyde  solution  (formalin)  to  3  cc.  of  pure  concentrated 
sulphuric  acid  just  before  using. 

Glinzburg's  Reagent.^ — Dissolve  i  part  of  phloroglucinol  and 
I  part  of  vanilline  in  30  parts  of  alcohol.  This  reagent  is  used 
to  detect  free  mineral  acid,  especially  hydrochloric  acid,  but  it 
does  not  react  with  free  organic  acids. 

Hiinef eld's  Solution.— Add  25  cc.  of  alcohol,  5  cc.  of  chloro- 
form and  1.5  cc.  of  galcial  acetic  acid  to  15  cc.  of  old  turpentine 
which  has  been  exposed  for  some  time  to  air  and  light.  The 
turpentine  used  should  not  produce  a  blue  color  with  guaiac 
tincture  direct  nor  with  1 5  cc.  of  3-5  per  cent,  hydrogen  peroxide 
free  from  acid.     This  solution  is  used  in  the  detection  of  blood. 

Iodic  Acid  Solution. — Prepare  a  10  percent,  aqueous  solution 
of  iodic  acid  (HIO3). 

Magnesia  Mixture. — Dissolve  11  grams  of  crystallized  mag- 
nesium chloride  (MgCl2.6H20)  and  14  grams  of  ammonium 
chloride  in  130  cc.  of  water  and  add  70  grams  of  ammonium 
hydroxide  solution  (sp.  gr.  0.96  =  10  per  cent,  of  NH3).  This 
mixture  should  be  clear.  It  is  used  to  detect  arsenic  and 
phosphoric  acids. 

Mandelin's  Reagent.^ — Dissolve  i  part  of  ammonium  meta- 
vanadate  (H4N.VO3)  in'200  parts  of  pure  concentrated  sulphuric 
acid. 

Millon's  Reagent.^ — Dissolve  i  part  of  mercury  in  i  part  of 
cold  fuming  nitric  acid.  Dilute  with  twice  the  volume  of 
water  and  decant  the  clear  solution  after  several  hours. 

Nessler's  Reagent. — Dissolve  separately  in  the  cold  3.5 
grams  of  potassium  iodide  in  10  cc.  of  water  and  1.7  grams  of 
mercuric  chloride  in  30  cc.  of  water.  Add  mercuric  chloride 
solution  to  potassium  iodide  solution  until  there  is  a  permanent 
precipitate.  Dilute  with  20  per  cent,  sodium  hydroxide  solution 
until  the  volume  is  100  cc.  Add  mercuric  chloride  solution, 
until  there  is  again  a  permanent  precipitate  and  let  the  solution 

^  It  is  advisable  to  prepare  this  reagent  as  required.  Keep  two  separate  alco- 
holic solutions  (i  :  15)  of  phloroglucinol  and  vanilline  and  mix  volume  for  volume 
as  needed.    Tr. 


PKEPA RATION  OF  REAGENTS  315 

settle.  Decant  the  clear  solution  and  keep  in  small  bottles  in 
the  dark.     This  reagent  improves  upon  standing. 

Mecke's  Reagent.' — Dissolve  0.5  gram  of  selenious  acid  in 
10  grams  of  pure  concentrated  sulphuric  acid. 

Stannous  Chloride  Solution. — Mix  5  j)arts  of  crystalHzed 
stannous  chloride  with  i  part  of  hydrochloric  acid  and  com- 
pletely saturate  with  dry  hydrochloric  acid  gas.  Let  this 
solution  settle  and  filter  through  asbestos.  It  is  a  pale, 
yellowish,  refractive  hquid  (sp.  gr.  at  least  1.9).  This  solution 
is  used  to  detect  arsenic  (Bettendorff's  Arsenic  TestJ. 

C.  The  Indicator  lodeosine 

lodeosine,  or  erythrosine,  C20H8I4O6,  is  a  tetra-iodo-fluoresceine,  formed  by 
treating  fluoresceine  with  iodine  and  having  the  formula: 

/C6Hl2(OH)\o 
C^CeHIaCOH)/ 
I  \C6H4.CO.O 

I I 

The  commercial  preparation  usually  contains  as  impurities  small  quantities  of 
substances  almost  insoluble  in  ether.  To  obtain  a  pure  product,^  dissolve  com- 
mercial lodeosine  in  aqueous  ether  and  extract  lodeosine  from  the  filtered  ether 
solution  by  means  of  dilute  sodium  hydroxide  solution.  Strong  sodium  hydrox- 
ide solution,  added  to  this  aqueous  alkaUne  solution,  precipitates  the  sodium  salt 
of  lodeosine.  Filter,  wash  with  cold  alcohol  and  crystaUize  from  hot  alcohol. 
Well  formed,  almost  rectangular  plates  having  a  green  color  on  the  surface  are 
obtained.  Hydrochloric  acid  precipitates  pure  lodeosine  from  the  aqueous  solu- 
tion of  the  sodium  salt.  Pure  lodeosine  dried  at  1 20°  is  markedly  lighter  than  the 
commercial  preparation.  It  is  almost  insoluble  in  absolute  ether,  benzene  and 
chloroform;  more  easily  soluble  in  acetone,  alcohol  and  aqueous  ether.  The  tone 
of  the  purified  pigment  dissolved  in  aqueous  alkali  is  yellower  than  that  of  the 
crude  product.  lodeosine  is  a  scarlet  crystalline  powder  which  dissolves  in  alcohol 
with  a  deep  red  and  in  ether  with  a  yello\\'ish  red  color.  lodeosine  is  said  to  be 
insoluble  in  water  containing  a  trace  of  hydrochloric  acid.  To  prepare  lodeosine 
solution  for  use  as  an  indicator,  dissolve  i  gram  of  the  pigment  in  500  grams  of 
alcohol. 

^  Zeitschrift  fiir  offentHche  Chemie  5,  350  (1899). 

2  Fr.  Mylius  and  F.  Foerster,  Berichte  der  Deutschen  chemischen  Gesellschaft 
24,  1482  (1891). 


INDEX 


Abrin,  221 

Absorption  spectra,  299 

Acetanilide,  68 

Acetone,  51 

Acid,  cacodylic,  238 

,  carbolic,  26 

,  with  aniline,  34 

,  hydrochloric,  176 

,  hydrocyanic,  19 

,  with   potassium   ferrocyan- 

ide,  25 

,  hypophosphorous,  8 

■ -,  iodic,  reagent,  314 

,  meconic,  205 

,  nitric,  177 

,  oxalic,  182 

,  phospho-molybdic,  reagent,  312 

,  phosphorous,  14 

,  phosphoric,  7 

,  phospho-tungstic,  reagent,  312 

,  picric,  6s 

,  reagent,  313 

,  picrolonic,  reagent,  313 

,  saUcylic,  72 

,  in  foods  and  beverages,  243 


,  selenious,  reagent,  207 

,  sulphuric,  180 

,  sulphurous,  181 

,  tannic,  reagent,  312 

Acids,  mineral,  175 

Aconitine,  estimation  in  aconite  root, 

255 
Alcohol,  ethyl,  49 
AlkaHes,  186 

Alkaloids,  Stas-Otto  method,  59 
Aloin,  reagent,  307 
Aluminium  acetate,  reagent,  282 
Ammonia,  185 
AniUne,  44,  89 


Antimony,  157 

,  fate,  distribution  and  elimina- 
tion, 168 

,  mirror  and  spot,  153 

,  quantitative  determination,  234 

Antipyrine,  78,  118 

Apomorphine,  122 

Arrhenal,  239 

Arsenic,  Marsh-Berzelius  test,  149 

,  Bettendorff's  test,  155 

,  biological  test,  235 

,  bulb-tube  test,  155 

,  detection,  149 

,  distinction  from  antimony,  153 

,  electrolytic  detection,  226,  230 

,  fate,  distribution  and  elimina- 
tion, 166 

,  Fresenius-v.  Babo  test,  154 

,  Gutzeit  test,  156,  233 

,  in  organic  compounds,  238 

,  in  presence   of    organic    matter, 

226 

,  isolation  as  trichloride,  226 

,  minute  amounts,  240 

,  mirror  and  spot,  153 

,  normal,  167 

Assajdng  of  alkaloids  by  E.  Merck,  295 

Atoxyl,  239 

Atropa  belladonna,  estimation  of  alka- 
loids, 293 

Atropine,  xoo 

,  estimation,  293 

Barium,  164 
Benzaldehyde,  53 
Berberine,  estimation  of,  275 
Bettendorff's  arsenic  test,  155 
Biological  arsenic  test,  235 
Biological  blood  test,  307 


317 


318 


INDEX 


Bismuth,  i6o 

,  i&te,    distribution,    elimination, 

173 
Bitter  almond  water,  53 
Blondlot-Dusart  test  for  phosphorus,  8 
Blood,  biological  test,  307 

,  carbon  monoxide,  297 

,  coagulation,  222 

,  defibrinated,  222 

,  spectroscopic  test,  303 

,  tests,  305 

Blood-stains,  300 

Brucine,  96 

,  estimation  in  nux  vomica,  286 


Cadmium,  160 

Caffeine,  79,  118 

,  estimation   in    coffee,    tea    and 

cola-nuts,  264 

,  estimation  in  cacao  and  choco- 
late, 291 

Cantharidin,  196 

,  estimation  in  Spanish  flies,  256 

Carbolic  acid,  26 

Carbon  disulphide,  42 

,  estimation  in  air,  48 

Carbon  monoxide  in  blood,  247 

Carboxy-hsemoglobin,  absorption-spec- 
trum, 299 

Carmine,  absorption-spectrum,  304 

Cephasline,  estimation  in  ipecac,  271 

Chavicine,  280 

Chloral  hydrate,  38 

,  as  a  solvent,  244 

Chloroform,  35 

,  estimation  in  cadavers,  37 

Choline,  204 

Chromium,  162 

,  fate,  distribution  and  elimina- 
tion, 170 

Cinchona  alkaloids,  estimation  in  bark, 
261 

Cinchonidine,  257 

Cinchonine,  257 

Cocaine,  loi 

Codeine,  106 

,  estimation  as  picrolonate,  247 


Colchicin,  64 

,  estimation  in  seed  and  corms, 

262 

Coniine,  85 

Copper,  157 

,  fate,  distribution  and  elimina- 
tion, 171 

Crotin,  222 

Cytisine,  198 

Destruction  of  organic  matter,  141 

Digitalin,  201 

Digitahs  glucosides,  200 

Digitonin,  200 

Digitoxin,  200 

Distillation,  for  phosphorus,  5 

,  for  volatile  poisons,  18 

Emetine,  270 
Erdmann's  reagent,  313 
Ergot,  202 

Ergotinine,  estimation,  204 
Eserine,  105 
Ethyl  alcohol,  49 

Extract  of  belladonna,  estimation,  248, 
294,  295 

of  cinchona,  estimation,  259 

of  hyoscyamus,  estimation,  294 

of  opium,  estimation,  278 

of  nux  vomica,  estimation,  288 

Fehling's  solution,  313 
Formaldehyde-sulphuric  acid,  reagent, 

314 
Fresenius-v.  Babo  apparatus,  154 
Froehde's  reagent,  313 
Fuchsine,  absorption  spectrum,  305 

General  alkaloidal  reagents,  310 
Githagin,  216 

Gold  chloride,  reagent,  310 
Guaiac,  chloral  hydrate  solution,  rea- 
gent, 305 
Guaiac-copper  paper,  21 
Giinzburg's  reagent,  314 
Gutzeit's  arsenic  test,  156,  233 

Hsematin,  303 


INDEX 


319 


Haematoporphyrin ,   absorption-spec- 
trum, 299 

,  in  urine,  195 

Haemin  crystals,  301 

Hajmocliromogen,        absorption  -  spec 
trum,  303 

Hemoglobin,  absorption-s  p  e  c  t  r  u  m, 
299 

Haemolysis,  216 

,  toxicity  estimated  by,  251 

Homatropine,  loi 

Human  blood,  detection,  305 

Hunef eld's  solution,  314 

Hydrastine,  112 

,  estimation,  274 

Hydrastinine,  113 

Hydrochloric  acid,  176 

Hydrocyanic  acid,  19 

Hydrogen  sulphide,  arsenic-free,  145 

Hyoscyamine,  99 

lodeosine,  315 
Iodic  acid,  reagent,  314 
Iodoform,  41 

lodo-potassium  iodide,   estimation   of 
alkaloids,  250 

,  reagent,  311 

Ipecac,  estimation  of  alkaloid  in,  270 
Isopelletierine,  263 

Lead,  160,  164 

,  fate,   distribution   and   elimina- 
tion, 169 
Lead  paper,  test  for  phosphorus,  3 

Magnesia  mixture,  reagent,  7,  314 
Maltol,  244 

Mandelin's  reagent,  314 
Marsh-Berzelius  apparatus,  151 
Mecke's  reagent,  315 
Meconic  acid,  205 
Meconine,  206 

Mercuric  chloride,  reagent,  311 
- — — ,  cyanide,  25 
Mercury,  158 

— — ,  fate,    distribution    and  elimina- 
tion, 171 
Metallic  poisons,  141 


Metals,  distribution  and  cUmination, 

165 
Methsemoglobin ,  absorption-spectrum, 

299 
Milk,  salicylic  acid  in,  243 
Millon's  reagent,  314 
Mineral  acids,  175 
Mitscherlich  apparatus,  5 
Morphine,  126 
— — ,  estimation,  247,  276 

Narceine,  131 
Narcotine,  108 
Nessler's  reagent,  314 
Nicotine,  86 

,  estimation  in  tobacco,  272 

Nitric  acid,  177 

Nitrobenzene,  42 

Non-volatile  organic  poisons,  57 

Opium,  205 
Oxalic  acid,  182 

Oxyhjemoglobin,  absorption-spectrum, 
299 

Papaverine,  208 

Paraxanthine,  293 

Pelletierine,  263 

Phenacetine,  70 

Phenol,  26 

Phospho-molybdic  acid,  reagent,  312 

Phosphorous  acid,  14 

Phosphorus,  5 

■,  Blondlot-Dusart  test,  8 

,  estimation,  15 

,  in  oils,  14,  224 

,  Hilger-Nattermann,  11 

,  ]\Iitscherhch,  5 

Phospho-tungstic  acid,  reagent,  312 
Physiological  salt  solution,  216 

•  test  for  atropine,  100 

• for  cantharidin,  198 

for  cocaine,  105 

for  phj-sostigmine,  106 

for  strychnine,  95 


Physostigmine,  105 
Picraconitine,  254 


320 


INDEX 


Picric  acid,  65 

,  reagent,  313 

Picrotoxin,  61   ' 
Pilocarpine,  210 

,  estimation,  279 

Piperidine,  280 
Piperine,  280 

Platinum  chloride,  reagent,  311 
Pomegranate  bark,  alkaloids  in,  263 
Potassium    bismuthous    iodide,    esti- 
mation of  alkaloids,  248 

,  reagent,  311 

■  cadmium  iodide,  reagent,  311 

chlorate,  187 

,  to  destroy  organic  matter,  141 


mercuric  iodide,  reagent,  311 

zinc  iodide,  reagent,  312 

Pseudo-pelletierine,  263 
Psychotrine,  276 
Ptomaines,  212 
Pyramidone,  119 

Quinidine,  257 

Quinine,  114 

,  estimation,  251,  261 

Ricin,  221 


Solanidine,  218 
Solanine,  217 

,  estimation,  284 

Stannous  chloride,  reagent,  315 
Stas-Otto  process,  59 
Stypticine,  estimation,  246 
Strychnine,  92 

,  estimation  with  quinine,  251 

,  estimation  in  nux  vomica,  291 

Sulphonal,  193 
Synopsis  of  Group  I,  SS 

n,  134 

Ill,  164 

Tannic  acid,  reagent,  312 
Teichmann's  crystals,  302 
Tellurium,  biological  arsenic  test,  236 
Thebaine,  220 
Theine  (see  Caffeine). 
Theobromine,  estimation  in  cacao,  291 
TheophylUne,  293 
Tin,  157 

,  fate,   distribution  and  eUmina- 

tion,  174 
Toxalbumins,  221 
Trional,  196 


SalicyUc  acid,  72 
Santonin,  192 

,  estimation  in  wormseed,  282 

,  estimation  in  troches,  284 

Saponins,  213 

Schaer's  blood  tests,  306,  308 
Scherer's  phosphorus  test,  3 
Schlererythrin,  203 
Schonbein-Van  Deen  blood  test,  305 
Selenious  acid,  reagent,  207 
Selenium,  biological  arsenic  test,  236 
Silver,  164 

,  fate,   distribution  and  ehmina- 

tion,  172 


Uranium,  173 

Van  Deen's  blood  test,  305 
Veratrine,  89 
Veronal,  75 
Volatile  poisons,  3 

Wine,  salicylic  acid  in,  243 

Zinc,  161 

,  fate,  distribution  and  elimina- 
tion, 173 


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